Human Infectiology MDx | Catalogue 2011 - Diagenode Diagnostics

The secure way to performanceCATALOGUE 2011 CONTENTSCustomized test 4The Universal QC Standard 4The Technology of Diagenode’s IVD Kits 5PCR ControlsDNA Extraction & PCR Inhibition Control 6RNA Extraction & PCR Inhibition Control 6Universal Inhibition Control 7Gastroenteritis infectonsBacteriaClostridium difficile 8Gastroenteritis bacteria multiplex 9Shigella 10ParasitesDientamoeba fragilis 11Gastroenteritis parasites multiplex 12VirusesGastroenteritis viruses multiplex 13Norovirus I & II 14Sexually transmitted infectonsChlamydia trachomatis 15Chlamydia trachomatis & Neisseria gonorrhoeae 16Mycoplasma genitalium 17Neisseria gonorrhoeae 18Trichomonas vaginalis 19Herpes virusesCytomegalovirus 20Epstein-Barr virus 21Herpesvirus six 22Herpes simplex 1 / Herpes simplex 2 23Varicella zoster virus 24Respiratory infectonsBordetella 25Bordetella New multiplex 26Human influenza A/B 27Human metapneumovirus 28Human Parainfluenza 1 / 3 29Human Parainfluenza 2 / 4 30Human respiratory syncytial virus 31Human rhinovirus 32Legionella 33Mycobacterium tuberculosis 34Mycoplasma pneumoniae & Chlamydophila pneumoniae 35Respiratory RNA viruses multiplex 36Swine Influenza 37MeningitisNeisseria meningitis / Serogroup B & C 38Neisseria meningitis / Serogroup A, B, C, W135 & Y 39Neisseria meningitidis & Steptococcus pneumoniae 40Other pathogensAdenovirus 41Borrelia burgdorferi 42Enterovirus 43Parvovirus B19 443

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011Customized testCustomized test for Diagnostic laboratoriesDiagenode is a leading company in the In vitro Diagnostics industry. We develop, manufacture,and market state-of-the-art qPCR or LAMP test kits for infectious diseases.In addition to our wide portfolio of off-the-shelf IVD kits, we provide customized tests uponrequest.Diagenode’s proprietary knowledge, in-house expertise, and strong relationship with leadingscientists and medical professionals enable us to provide quality customized kits for yourspecific testing needs. Our kits can be optimized for nearly any application and always meetfull regulatory requirements.The Universal QC Standard“The Universal Standard” for Real Time PCR Pathogen IVD Quality ControlOur kits utilize smart design and include universal, validated controls with every kit for highestaccuracy. These unique kit controls are compatible with all DNA amplification-based pathogendetection methods and with most nucleic acid extraction methods and tools. An increasingnumber of Diagenode’s customers are additionaly using the controls for the validation (CEmarking) of their own homebrew reagents or with kits from other suppliers.We offer a variety of controls which are labeled with the most commonly used dyes : YellowDye, Orange Dye, Texas Red, and C5 Dye, allowing flexibility of use with any qPCR machine.Customization options include:• Expansion of best primer pairs/probes combinations• Optimization of RT-PCR simplex and multiplex conditions• Expansion of control offerings• CE certification and clinical studies• Bulk production of any kitOur products are manufactured in facilities that are GMP compliant.Norm. Fluoro0.60.50.40.30.20.1Results of DNA extraction & PCR inhibition controlwith Cytomegalovirus kit (dia-CMVQ-050)QCMD08 CMV sampleStandard curve Dia-CMVQ-0500.0510 15 20 25Cycle30 35 40 45See page:6 for DNA extraction & PCR inhibition control, ref # Dia-EIC/DNA-050 and for RNA extraction & PCRinhibition control, ref # Dia-EIC/RNA-050.7 for Universal inhibition control, ref # Dia-UIC/DNA-050.4 | www.diagenodediagnostics.com

The Technology of Diagenode’s IVD KitsTARGED DNA (DOUBLE STRANDED)Real-time polymerase chain reaction, also called quantitative real-time polymerase chainreaction (qPCR), is a nucleic acid technology used to detect and quantify specific DNAsequences. This quantification can be measured in absolute or relative number of copies.Real-time PCR technology allows for rapid, specific, and quantitative measurements of thepresence of genes involved in infectious diseases, cancer, and genetic abnormalities. Thispowerful technology is utilized in Diagenode’s kits for accurate pathogen detection for anumber of diseases.REPORTER AND QUENCHER-MARKEDPROBETAQ POLYMERASEOne of the prominent attributes of qPCR technology is that amplified DNA is quantified asit accumulates in the reaction in “real-time” for each PCR amplification cycle, allowingfor far more accurate measurements than with traditional end-point PCR. These realtimemeasurements are made possible by quantifying signals from fluorescent dyes thatintercalate with the double-stranded DNA produced in each amplification cycle. Alternatively,one can use labeled DNA oligonucleotide probes with a fluorophore (e.g. a TaqMan ® probe)that subsequently hybridize with the complementary DNA to allow real-time measurementsduring qPCR. The resulting fluorescence signals can be closely monitored and quantifiedusing an instrument such as a thermal cycler adapted for qPCR.REPORTER AND QUENCHER-MARKEDPROBETARGED DNA (DOUBLE STRANDED)RELEASED REPORTER TARGED DNA STRAND WITHFLUOROPHORE HYBRIDIZED PROBETAQ POLYMERASEMultiplex assaysTARGED DNA (DOUBLE STRANDED)RELEASED REPORTERFLUOROPHOREThe ability to multiplex PCR adds power to real-time analysis. Multiplex real-time PCRcombines multiple PCR assays together into a single reaction to conserve precious samplematerial, reduce contamination, reduce handling of a large number of individual reactions,and save time. The DNA targets are amplified simultaneously yet detected independently withdifferent reporters (fluorescent probes labeled with FAM, VIC, NED, Texas Red, and other dyes)with distinct wavelengths. Most qPCR platforms have the ability to measure the spectra ofthese commonly used dyes. Multiplex assays also allow samples and internal controls to beco-amplified with their targets and thus permit specific and sensitive discrimination in singletube. Diagenode utilizes multiplex qPCR to deliver better patient care with:• Fast results without delay - Analysis of up to 4 targets in a single reaction• High sensitivity - Sensitive detection of as few as 1-10 copies of each target• Reliability - Quantification of target and reference genes in the same well• Cost-effectiveness – Multiple assays conducted in a single reactionReferencesTARGED DNA STRAND WITHHYBRIDIZED PROBE1. Nailis H, Coenye T, Van Nieuwerburgh F, Deforce D, Nelis HJ (2006). «Development and evaluation ofdifferent normalization strategies for gene expression studies in Candida albicans biofilms by real-timePCR». BMC Mol Biol. 7: 25. PMID 16889665.2. Nolan T, Hands RE, Bustin SA (2006). «Quantification of mRNA using real-time RT-PCR». Nat. Protoc. 1:1559-1582. PMID 17406449.TARGED DNA STRAND WITHHYBRIDIZED PROBE5

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011PCR CONTROLSDNA extraction & PCRinhibition controlRef: Dia-EIC/DNA-050DNA extraction & inhibition control is a DNA virus showingno sequence identity with human genome, viruses and otherpathogens affecting human being.DNA extraction & inhibition control is a complete virus, havingpreserved its infectious power.This virus is listed in the class Ea1 established by the EuropeanFederation of Biotechnologies (EFP). This virus is not reported asbeing dangerous for human being.RNA extraction & PCRinhibition controlRef: Dia-EIC/RNA-050RNA extraction & inhibition control is a RNA virus showingno sequence identity with human genome, viruses and otherpathogens affecting human being.RNA extraction & inhibition control is a complete virus, havingpreserved its infectious power.This virus is listed in the class Ea1 established by the EuropeanFederation of Biotechnologies (EFP). This virus is not reported asbeing dangerous for human beingVALIDATIONNumber of wells by test: 1 (with the pathogen researched)Samples used:Samples validated for all diagenode pathogen detection kits.Validated extraction methods: Extraction, DNA, validated for all diagenode pathogen detection kits.Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 50 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)Dyes available:• DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nm• DIA-EIC/DNA (DR)-050: orange dye, emission: 576 nm• DIA-EIC/DNA (TR)-050: texas red, emission: 603 nm• DIA-EIC/DNA (Cy5)-050: Cy5 dye, emission: 662 nmNumber of wells by test: 1 (with the pathogen researched)Samples used:Samples validated for all diagenode pathogen detection kits.Validated extraction methods: Extraction, RNA, validated for all diagenode pathogen detection kits.Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 50 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)Dyes available:• DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nm• DIA-EIC/RNA (DR)-050: orange dye, emission: 576 nm• DIA-EIC/RNA (TR)-050: texas red, emission: 603 nm• DIA-EIC/RNA (Cy5)-050: Cy5 dye, emission: 662 nm6 | www.diagenodediagnostics.com

Universal inhibition controlRef: Dia-UIC-050This product (linearized plasmid) consists of an inhibition controlfor real-time PCR reactions.VALIDATIONNumber of wells by test: 1 (with the pathogen researched)Samples used:Samples validated for all diagenode pathogen detection kits.Validated extraction methods: Extraction, DNA & RNA, validated for all diagenode pathogen detection kits.Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 50 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)Dyes available:• DIA-UIC(YD)-050: yellow dye, emission: 549 nm• DIA-UIC(DR)-050: orange dye, emission: 576 nm• DIA-UIC(TR)-050: texas red, emission: 603 nm• DIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nmPerfect compatibility with most regular Real-Time PCR systemsABI 7000/7300/7900 * ABI 7500 * Roche Lc480 Roche Lc2.0Bio-rad ICyclerIQ5/CFX96Qiagen *Rotor-GeneStratageneMx3000/3005PCepheid SmartCycler IIDiagenode DyesDIA-EIC/DNA(YD)-050DIA-EIC/RNA(YD)-050DIA-UIC(YD)-050DIA-EIC/DNA(DR)-050DIA-EIC/RNA(DR)-050DIA-UIC(DR)-050DIA-EIC/DNA(TR)-050DIA-EIC/RNA(TR)-050DIA-UIC(TR)-050DIA-EIC/DNA(Cy5)-050DIA-EIC/RNA(Cy5)-050DIA-UIC(Cy5)-050VIC VIC VIC* VIC* HEX Green channel FAM Alexa 532NEDNEDTexas red Texas red * Texas red / ROX Orange channel ROX Texas redCy5 Cy5 Cy5 Red channel Cy5 Alexa 647 *** ABI systems: for quencher, select None. Roche systems: compensation file (ccc) must be done. Qiagen Rotor-Gene system: perform optimisation at 60°C at beginning of run.** Coming soon for Alexa 647: DIA-EIC/DNA(RD)-050 / DIA-EIC/RNA(RD)-050 / DIA-UIC(RD)-0507

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011GASTROENTERITIS INFECTIONSClostridium difficileRef: Dia-Cdiff-050Clostridium difficile is a species of Gram-positive bacteria ofthe genus Clostridium that causes diarrhea and other intestinaldisease when competing bacteria are wiped out by antibiotics.C. difficile is the most serious cause of antibiotic-associateddiarrhea (AAD) and can lead to pseudomembranous colitis, asevere infection of the colon, often resulting from eradicationof the normal gut flora by antibiotics (1) . The C. difficile bacteria,which naturally reside in the body, become overpopulated: Theoverpopulation is harmful because the bacterium releasestoxins that can cause bloating, constipation, and diarrhea withabdominal pain, which may become severe. Latent symptomsoften mimic some flu-like symptoms. Often, it can be curedsimply by discontinuing the antibiotics responsible (2) .(1)Curry J (2007-07-20). “Pseudomembranous Colitis”. eMedicine. WebMD.http://www.emedicine.com/med/topic1942.htm#section~introduction.Retrieved 2008-11-17. (2) Ryan KJ, Ray CG (editors) (2004). Sherris MedicalMicrobiology (4th ed.). McGraw Hill. pp. 322–4.VALIDATIONResearched pathogens:• Clostridium difficile, tcdA: FAM dye, emission: 520 nm• Clostridium difficile, tcdB: yellow dye, emission: 553 nmTarget genes: • tcdA (enterotoxine A)• tcdB (enterotoxine B)Number of wells by test: 1 (Clostridium difficile enterotoxin A / Clostridium difficile enterotoxin B)Samples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche lightCycler (480/2,0 on request)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR, final volume)• 100 (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmDIA-EIC/DNA(RD)-050: Red dye, emission: 669 nm8 | www.diagenodediagnostics.com

Gastroenteritisbacteria multiplexRef: Dia-GE/bacteria.I-050Gastroenteritis/bacteria panel I allows the detection of Salmonellaenterica and Campylobacter jejuni.Salmonella enterica (formerly Salmonella choleraesuis) is a rodshaped, flagellated, aerobic, Gram-negative bacterium, and amember of the genus Salmonella. S. enterica often infects cattleand poultry, though also other animals such as domestic catsand hamsters have also been shown to be sources of infection tohumans. However, investigations of vacuum cleaner bags haveshown that households can act as a reservoir of the bacterium;this is more likely if the household has contact with an infectionsource, for example members working with cattle or in a veterinaryclinic.Campylobacter jejuni is a species of curved, rod-shaped, nonsporeforming, Gram-negative microaerophilic, bacteria commonlyfound in animal feces. It is one of the most common causes ofhuman gastroenteritis in the world. Food poisoning caused byCampylobacter species can be severely debilitating but is rarelylife-threatening. It has been linked with subsequent developmentof Guillain-Barré syndrome (GBS), which usually develops two tothree weeks after the initial illness. Infection with C. jejuni usuallyresults in enteritis, which is characterised by abdominal pain,diarrhea, fever, and malaise. The symptoms usually persist forbetween 24 hours and a week, but may be longer. Diarrhea canvary in severity from loose stools to bloody stools.VALIDATIONResearched pathogen:Target genes:• Salmonella enterica : FAM dye, emission: 520 nm• Campylobacter jejuni : yellow dye, emission: 553 nm• invA (Salmonella enterica)• mapA (Campylobacter jejuni)Number of wells by test: 1 (Salmonella enterica / Campylobacter jejuni)Samples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche lightCycler (480/2,0 on request)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR, final volume)• 100 (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm9

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011GASTROENTERITIS INFECTIONSShigellaRef: Dia-Shigella-050Shigella is a genus of Gram-negative, non-spore formingrod-shaped bacteria closely related to Escherichia coli andSalmonella.Shigella species are classified by four serogroups:• Serogroup A: S. dysenteriae (12 serotypes)• Serogroup B: S. flexneri (6 serotypes)• Serogroup C: S. boydii (23 serotypes)• Serogroup D: S. sonnei (1 serotype)Shigella causes dysentery that results in the destruction of theepithelial cells of the intestinal mucosa in the cecum and rectum.Some strains produce enterotoxin and Shiga toxin, similar to theverotoxin of E. coli O157:H7 (1) . Both Shiga toxin and verotoxin areassociated with causing hemolytic uremic syndrome.(1)Hale TL, Keusch GT (1996). Shigella. in: Baron’s Medical Microbiology (BaronS et al., eds.) (4th ed.). Univ of Texas Medical Branch.VALIDATIONResearched pathogen:• Shigella: FAM dye, emission: 520 nmTarget genes:• ipaH (Shigella)Number of wells by test: 1 (Shigella)Samples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche lightCycler (2.0/480)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR)• 100 (25 µl PCR)• 120 (20 µl PCR)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm10 | www.diagenodediagnostics.com

Dientamoeba fragilisRef: Dia-DF-050Dientamoeba fragilis is a single celled parasite, despite its namedientamoeba fragilis its not an amoeba but flagellate found inthe gastrointestinal tract of some humans, pigs and gorillas. Insome people it causes gastrointestinal upset while in others itdoes not (1) It is an important cause of traveler’s diarrhea, chronicdiarrhea, abdominal cramping and loss of appetite.(1)Windsor JJ, Macfarlane L (May 2005). “Irritable bowel syndrome: the needto exclude Dientamoeba fragilis”. Am. J. Trop. Med. Hyg. 72 (5): 501; authorreply 501–2. PMID 15891119. http://www.ajtmh.org/cgi/pmidlookup?view=long&pmid=15891119.J.J. Verweij et al., Real-time PCR for the detection of Dientamoeba fragilis infecal samples, Molecular and Cellular Probes 21 (2007), p. 400-404VALIDATIONResearched pathogen:Dientamoeba fragilis: FAM dye, emission: 520 nmTarget genes:5.8S rRNANumber of wells by test: 1 (Dientamoeba fragilis)Samples used:StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7500/7900)• Roche lightCycler (480)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR, final volume)• 100 (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm11

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011GASTROENTERITIS INFECTIONSGastroenteritisparasites multiplexRef: Dia-GE/Parasite.I-050Gastroenteritis/Parasite panel I allows the detection of Entamoebahistolytica, Giardia intestinalis and Cryptosporidium parvum.Entamoeba histolytica is an anaerobic parasitic protozoan, partof the genus Entamoeba. Infection can lead to amoebic dysenteryor amoebic liver abscess. Symptoms can include fulminatingdysentery, bloody diarrhea, weight loss, fatigue, abdominal pain,and amoeboma. The amoeba can actually bore into the intestinalwall, causing lesions and intestinal symptoms, and it may reachthe blood stream (1) .Giardia intestinalis is a flagellated protozoan parasite thatcolonises and reproduces in the small intestine, causing giardiasis.The giardia parasite attaches to the epithelium by a ventraladhesive disc, and reproduces via binary fission (2) . Giardiasis doesnot spread via the bloodstream, nor does it spread to other partsof the gastro-intestinal tract, but remains confined to the lumenof the small intestine (3) .Cryptosporidium parvum is one of several species that causecryptosporidiosis, a parasitic disease of the mammalian intestinaltract. Primary symptoms of C. parvum infection are acute, watery,and non-bloody diarrhoea. C. parvum infection is of particularconcern in immunocompromised patients, where diarrhea canreach 10–15L per day. Other symptoms may include anorexia,nausea/vomiting and abdominal pain (4) .VALIDATIONResearched pathogen:Target genes:• Entamoeba histolytica: FAM dye, emission: 520 nm• Giardia intestinalis: yellow dye, emission: 553 nm• Cryptosporidium parvum: texas red dye, emission: 603 nm• 18S rRNA (Entamoeba histolytica)• 8S rRNA (Giardia intestinalis)• seqment A (Cryptosporidium parvum)Number of wells by test: 1 (Entamoeba histolytica / Giardia intestinalis / Cryptosporidium parvum)Samples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7500/7900)• Roche lightCycler (480)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR, final volume)• 100 (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm(1)Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed.). McGraw Hill. pp. 733–8.(2)Oxford textbook of Medicine, Fourth Edition, Volume 1. Oxford University Press pp759-760.(3)Harrison’s Internal Medicine, Harrison’s Online Chapter 199 Protozoal intestinal infections and trochomoniasis.(4)”Cryptosporidiosis.” Laboratory Identification of Parasites of Public Health Concern. CDC. 5 Sept 2007.12 | www.diagenodediagnostics.com

Gastroenteritisviruses multiplexRef: Dia-GE/Virus.I-050Gastroenteritis/virus panel I allows the detection of Astrovirus,Rotavirus A, Norovirus 1 and 2 and Adenovirus.Astrovirus has a non-segmented, single stranded, positivesense RNA genome within a non-enveloped icosahedral capsid (1) .Astroviruses are now recognised as a cause of gastroenteritis inchildren and adults. The main symptoms are diarrhea, followedby nausea, vomiting, fever, malaise and abdominal pain. Someresearch studies have shown that the duration of the symptomsare approximately three to four days. Astrovirus infection is notusually a severe situation and only in some rare cases leads todehydrationRotavirus is the most common cause of severe diarrhea amonginfants and young children (2) , and is one of several viruses thatcause infections often called stomach flu, despite having norelation to influenza. It is a genus of double-stranded RNA virus inthe family Reoviridae. By the age of five, nearly every child in theworld has been infected with rotavirus at least once (3) . However,with each infection, immunity develops, subsequent infectionsare less severe (4) , and adults are rarely affected (5) . There are fivespecies of this virus, referred to as A, B, C, D, and E. RotavirusA, the most common, causes more than 90% of infections inhumans.Norovirus is an RNA virus that causes approximately 90% ofepidemic non-bacterial outbreaks of gastroenteritis around theworld (6) , and may be responsible for 50% of all foodborne outbreaksof gastroenteritis in the US (7) . Norovirus affects people of all ages.The viruses are transmitted by faecally contaminated food orwater, by person-to-person contact (8) and via aerosolization of thevirus and subsequent contamination of surfaces (9) .Adenovirus is a nonenveloped icosahedral virus composed ofa nucleocapsid and a double-stranded linear DNA genome.There are 53 described serotypes in humans and gastroenteritisis mainly caused by serotypes 40 and 41. Some people withadenovirus gastroenteritis may shed the virus in their stools formonths after getting over the symptoms.VALIDATIONResearched pathogen:Target genes:• Astrovirus: FAM dye, emission: 520 nm• Norovirus 1: Yellow dye, emission: 553 nm• Norovirus 2: Yellow dye, emission: 553 nm• Rotavirus A: Texas red dye, emission: 603 nm• Adenovirus: FAM dye, emission: 520 nm(RNA extraction & PCR inhibition control: Cy5, emission: 662 nm)(DNA extraction & PCR inhibition control: Cy5, emission: 662 nm)• ORF1a (Astrovirus)• RdRp/Capsid junction (Norovirus 1)• RdRp/Capsid junction) (Norovirus 2)• NSP3 (Rotavirus A)• Hexon (Adenovirus)Number of wells by test: 21. Astrovirus / Norovirus 1 / Rotavirus A / RNA extraction & PCR inhibition control2. Adenovirus / Norovirus 2 / DNA extraction & PCR inhibition controlSamples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7500/7900)• Roche lightCycler (480)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR)• 100 (25 µl PCR)(1)Matsui SM, Kiang D, Ginzton N, Chew T, Geigenmüller-Gnirke U (2001). «Molecular biology of astroviruses: selected highlights». NovartisFound. Symp. 238: 219–33; discussion 233–6. (2) Dennehy PH (2000). «Transmission of rotavirus and other enteric pathogens in the home».Pediatr. Infect. Dis. J. 19 (10 Suppl): S103–5. (3) Velázquez FR, Matson DO, Calva JJ, Guerrero L, Morrow AL, Carter-Campbell S, Glass RI, EstesMK, Pickering LK, Ruiz-Palacios GM (1996). «Rotavirus infections in infants as protection against subsequent infections». N. Engl. J. Med. 335(14): 1022–8. (4) Linhares AC, Gabbay YB, Mascarenhas JD, Freitas RB, Flewett TH, Beards GM (1988). «Epidemiology of rotavirus subgroups andserotypes in Belem, Brazil: a three-year study». Ann. Inst. Pasteur Virol. 139 (1): 89–99. (5) Bishop RF (1996). «Natural history of human rotavirusinfection». Arch. Virol. Suppl. 12: 119–28. (6) Lindesmith L, Moe C, Marionneau S, et al. (2003). «Human susceptibility and resistance to Norwalkvirus infection». Nat. Med. 9 (5): 548–53. (7) Widdowson MA, Sulka A, Bulens SN, et al. (2005). «Norovirus and foodborne disease, United States,1991-2000». Emerging Infect. Dis. 11 (1): 95–102. (8) Goodgame R (2006). «Norovirus gastroenteritis». Curr Gastroenterol Rep 8 (5): 401–8. (9) SaidMA, Perl TM, Sears CL (November 2008). «Healthcare epidemiology: gastrointestinal flu: norovirus in health care and long-term care facilities».Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America 47 (9): 1202–8.13

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011GASTROENTERITIS INFECTIONSNorovirus I & IIRef: Dia-Norovirus (I/II)-050Norovirus is an RNA virus that causes approximately 90% ofepidemic non-bacterial outbreaks of gastroenteritis around theworld (1) , and may be responsible for 50% of all foodborne outbreaksof gastroenteritis in the US (2) . Norovirus affects people of all ages.The viruses are transmitted by faecally contaminated food orwater, by person-to-person contact (3) and via aerosolization of thevirus and subsequent contamination of surfaces (4) .(1)Lindesmith L, Moe C, Marionneau S, et al. (2003). «Human susceptibility andresistance to Norwalk virus infection». Nat. Med. 9 (5): 548–53. (2) WiddowsonMA, Sulka A, Bulens SN, et al. (2005). «Norovirus and foodborne disease,United States, 1991-2000». Emerging Infect. Dis. 11 (1): 95–102. (3) GoodgameR (2006). «Norovirus gastroenteritis». Curr Gastroenterol Rep 8 (5): 401–8.(4)Said MA, Perl TM, Sears CL (November 2008). «Healthcare epidemiology:gastrointestinal flu: norovirus in health care and long-term care facilities».Clinical Infectious Diseases : an Official Publication of the Infectious DiseasesSociety of America 47 (9): 1202–8.VALIDATIONResearched pathogens:• Norovirus 1: FAM dye, emission: 520 nm• Norovirus 2: yellow dye, emission: 553 nmTarget genes: • RdRp/Capsid junction (Norovirus 1)• RdRp/Capsid junction) (Norovirus 2)Number of wells by test: 1 (Norovirus 1 / Norovirus 2)Samples used:• StoolValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Stool mini kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche lightCycler (480)• Biorad (ICycler/IQ5/CFX96)• Qiagen RotorGene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 (50 µl PCR)• 100 (25 µl PCR)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm14 | www.diagenodediagnostics.com

Chlamydia trachomatisRef: Dia-CT-050RUOSOONChlamydia trachomatis is a common sexually transmittedinfection (STI) in humans. C. trachomatis is only found inhumans (1) . Chlamydia is a major infectious cause of human genitaldisease. Chlamydia infection is one of the most common sexuallytransmitted infections worldwide (2) .C. trachomatis is naturally found living only inside human cells.Chlamydia can be transmitted during vaginal, anal, or oral sex,and can be passed from an infected mother to her baby duringvaginal childbirth. Between half and three-quarters of all womenwho have a chlamydia infection of the neck of the womb (cervicitis)have no symptoms and do not know that they are infected. Inmen, infection of the urethra (urethritis) is usually symptomatic,causing a white discharge from the penis with or without painon urinating (dysuria). Occasionally, the conditions spreads tothe upper genital tract in women (causing pelvic inflammatorydisease) or to the epididymis in men (causing epididymitis). Ifuntreated, chlamydial infections can cause serious reproductiveand other health problems with both short-term and long-termconsequences.(1)“www.chlamydiae.com (professional) - Taxonomy diagram”, http://www.chlamydiae.com/docs/Chlamydiales/diagram/taxondiag.htm, retrieved on2007-10-27. (2) Gerbase AC, Rowley JT, Mertens TE (1998), “Global epidemiologyof sexually transmitted diseases”, Lancet 351 Suppl 3: 2–4.VALIDATIONResearched pathogen:• Chlamydia trachomatis: FAM dye, emission: 520 nmTarget genes:• Cryptic plasmid (Chlamydia trachomatis)Number of wells by test: 1 (Chlamydia trachomatis)Samples used:• Urine• SwabValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• Amplicor CT/NG specimen preparation kit (CT/NG prep) (Roche)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmSEXUALLY TRANSMITTED INFECTIONS15

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011SEXUALLY TRANSMITTED INFECTIONSChlamydia trachomatis &Neisseria gonorrhoeaeRef: Dia-CT/NG-050RUOSOONChlamydia trachomatis is a common sexually transmittedinfection (STI) in humans. C. trachomatis is only found inhumans (1) . Chlamydia is a major infectious cause of human genitaldisease. Chlamydia infection is one of the most common sexuallytransmitted infections worldwide (2) .C. trachomatis is naturally found living only inside human cells.Chlamydia can be transmitted during vaginal, anal, or oral sex,and can be passed from an infected mother to her baby duringvaginal childbirth. Between half and three-quarters of all womenwho have a chlamydia infection of the neck of the womb (cervicitis)have no symptoms and do not know that they are infected. In men,infection of the urethra (arthritis) is usually symptomatic, causinga white discharge from the penis with or without pain on urinating(dysuria). Occasionally, the conditions spread to the upper genitaltract in women (causing pelvic inflammatory disease) or to theepididymis in men (causing epididymitis). If untreated, chlamydialinfections can cause serious reproductive and other healthproblems with both short-term and long-term consequences.Neisseria gonorrhoeae is a species of Gram-negative kidneybacteria responsible for the sexually transmitted diseasegonorrhoea (3) . Gonorrhea is an important public health problemand is the most common reportable infectious disease. Gonorrheais most frequently spread during sexual contact. However, it canalso be transmitted from the mother’s genital tract to the newbornduring birth, causing ophthalmia neonatorum and systemicneonatal infection. The incubation period is usually 2-8 days.In women, the cervix is the most common site of gonorrhea,resulting in endocervicitis and urethritis, which can be complicatedby pelvic inflammatory disease (PID). In men, gonorrhea causesanterior urethritis (4) .VALIDATIONResearched pathogens:Target genes:• Chlamydia trachomatis: FAM dye, emission: 520 nm• Neisseria gonorrhoeae: yellow dye, emission: 549 nm• Cryptic plasmid (Chlamydia trachomatis)• Opa, porA (Neisseria gonorrhoeae)Number of wells by test: 1 (Chlamydia trachomatis / Neisseria gonorrhoeae)Samples used:• Urine• SwabValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• Amplicor CT/NG specimen preparation kit (CT/NG prep) (Roche)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm(1)“www.chlamydiae.com (professional) - Taxonomy diagram”, http://www.chlamydiae.com/docs/Chlamydiales/diagram/taxondiag.htm, retrievedon 2007-10-27. (2) Gerbase AC, Rowley JT, Mertens TE (1998), “Global epidemiology of sexually transmitted diseases”, Lancet 351 Suppl 3: 2–4. (3)Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed. ed.). McGraw Hill. (4) Brian Wong, MD, Gonococcal Infections- emedicine,Apr 21, 2009 (http://emedicine.medscape.com/article/218059-overview)16 | www.diagenodediagnostics.com

Mycoplasma genatiliumRef: Dia-MG-050Mycoplasma genitalium is a small parasitic bacterium whichlives on the ciliated epithelial cells of the primate genital andrespiratory tracts.Infection by M. genitalium seems fairly common, can be transmittedbetween partners during unprotected sexual intercourse, and canbe treated with antibiotics.VALIDATIONResearched pathogen:• Mycoplasma genitalium: FAM dye, emission: 520 nmTarget genes:• adhesin (mgpB) (Mycoplasma genitalium)Number of wells by test: 1 (Mycoplasma genitalium)Samples used:• Urine• SwabValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm17

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011SEXUALLY TRANSMITTED INFECTIONSNeisseria gonorrhoeaeRef: Dia-NG-050Neisseria gonorrhoeae is a species of Gram-negative kidneybacteria responsible for the sexually transmitted diseasegonorrhoea (1) .Gonorrhea is a serious public health problem and is the mostcommonly reported infectious disease. Gonorrhea is mostfrequently spread during sexual contact. However, it can also betransmitted from the mother’s genital tract to the newborn duringbirth, causing ophthalmia neonatorum and systemic neonatalinfection. The incubation period is usually 2-8 days.In women, the cervix is the most common site of gonorrhea,resulting in endocervicitis and urethritis, which can be complicatedby pelvic inflammatory disease (PID). In men, gonorrhea causesanterior urethritis (2) .(1)Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed. ed.).McGraw Hill. (2) Brian Wong, MD, Gonococcal Infections- emedicine, Apr 21,2009 (http://emedicine.medscape.com/article/218059-overview)VALIDATIONResearched pathogen:• Neisseria gonorrhoeae: FAM dye, emission: 520 nmTarget genes:• opa, porA (Neisseria gonorrhoeae)Number of wells by test: 1 (Neisseria gonorrhoeae)Samples used:• Urine• SwabValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• Amplicor CT/NG specimen preparation kit (CT/NG prep) (Roche)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)• COBAS TaqMan 48PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm18 | www.diagenodediagnostics.com

Trichomonas vaginalisRef: Dia-TV-050Trichomonas vaginalis is an anaerobic, flagellated protozoan,a form of microorganism. The parasitic microorganism is thecausative agent of trichomoniasis, and is the most commonpathogenic protozoan infection of humans in industrializedcountries (1) . Infection rates between men and women are thesame with women showing symptoms while infections in men areusually asymptomatic. Transmission takes place directly becausethe trophozoite does not have a cyst product.(1)Soper D (January 2004). «Trichomoniasis: under control or undercontrolled?».American journal of obstetrics and gynecology 190 (1): 281–90VALIDATIONResearched pathogen:• Trichomonas vaginalis: FAM dye, emission: 520 nmTarget genes:• G3 hypothetical protein (Trichomonas vaginalis)Number of wells by test: 1 (Trichomonas vaginalis)Samples used:• Urine• SwabValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm19

Epstein-Barr VirusRef: Dia-EBVQ-050The Epstein-Barr Virus (EBV) is a virus of the herpes family, and isone of the most common viruses in humans. Most people becomeinfected with EBV at an early age, which is often asymptomaticbut may cause infectious mononucleosis. Symptoms of infectiousmononucleosis are fever, sore throat, and swollen lymph glands.Sometimes, a swollen spleen or liver involvement may develop.Heart problems or involvement of the central nervous systemoccurs only rarely, and infectious mononucleosis is almostnever fatal. There are no known associations between active EBVinfection and problems during pregnancy, such as miscarriage orbirth defects. Although the symptoms of infectious mononucleosisusually resolve in 1 or 2 months, EBV dormant or latent in afew cells in the throat and blood for life. Periodically, the viruscan reactivate and is commonly found in the saliva of infectedindividuals. This reactivation usually occurs without symptoms ofillness.Epstein-Barr Virus also establishes a lifelong dormant infectionin some cells of the body’s immune system. A late event in a veryfew carriers of this virus is the emergence of Burkitt’s lymphomaand nasopharyngeal carcinoma, two rare cancers that are notnormally found in the United States. EBV appears to play animportant role in these malignancies, but is probably not the solecause of disease.VALIDATIONResearched pathogen:• Epstein-Barr virus: FAM dye, emission: 520 nmTarget genes:• P143 (Epstein-Barr virus)Number of wells by test: 1 (Epstein-Barr virus) + 5 for the EBV quantitative curveSamples used:• Whole blood• PlasmaValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• QIAamp ® UltraSens Virus Kit (50) (Qiagen)• Maxwell 16 Blood DNA Purification kit (48) (Promega)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume) + 5 EBV quantitative curves• 100 reactions (25 µl PCR, final volume) + 5 EBV quantitative curves• 120 reactions (20 µl PCR, final volume) + 5 EBV quantitative curvesIn combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nm (included in the kit)DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm21

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011HERPES VIRUSESHerpesvirus SixRef: Dia-HHV6-050Human Herpesvirus Six (HHV-6) is one of the eight known virusesthat are members of the human herpervirus family. It causes thedisease Roseola.HHV-6 re-activation causes severe disease in transplant recipientsand can lead to graft rejection, often in consort with otherbetaherpesviridae. Likewise in HIV/AIDS, HHV-6 re-activationscause disseminated infections leading to end organ disease anddeath.HHV-6 has been reported in multiple sclerosis patients (1) andhas been implicated as a co-factor in several other diseases,including chronic fatigue syndrome (2), fibromyalgia, etc.(1)Alvarez-Lafuente R, Martín-Estefanía C, de las Heras V, et al. (February2002). “Prevalence of herpesvirus DNA in MS patients and healthy blooddonors”. Acta Neurol. Scand. 105 (2): 95–9. (2) Komaroff AL. (December 2006).“Is human herpesvirus-6 a trigger for chronic fatigue syndrome”. J Clin Virol.37 (Suppl 1): S39–46.VALIDATIONResearched pathogen:• Herpes 6: FAM dye, emission: 520 nmTarget genes: • ie gene (Herpes 6)Number of wells by test: 1 (Herpes 6) + 5 for the HHV 6 quantitative curveSamples used:• Serum• PlasmaValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system MagNa pure LC Total Nucleic Acid Isolation Kit(Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR) + 5 HHV 6 quantitative curves• 100 reactions (25 µl PCR) + 5 HHV 6 quantitative curves• 120 reactions (20 µl PCR) + 5 HHV 6 quantitative curvesIn combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nm (included in the kit)DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm22 | www.diagenodediagnostics.com

Herpes simplex 1 /Herpes simplex 2Ref: Dia-HSV1/2-050Herpes simplex viruses 1 and 2 (HSV-1 and HSV-2) are membersof the herpes virus family, Herpesviridae, which cause infectionsin humans (Ryan and Ray, 2004). All viruses in the herpes familyproduce life-long infections.These viruses, also known as Human Herpes Virus 1 and 2 (HHV-1and HHV-2) are neurotropic and neuroinvasive viruses; they enterand latent in the human nervous system, accounting for theirdurability in the human body. HSV-1 is commonly associated withherpes outbreaks of the face known as cold sores or fever blisters,whereas HSV-2 is more often associated with genital herpes.Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed. ed.).McGraw Hill. pp. 555–62.VALIDATIONResearched pathogens:• Herpes simplex virus 1: FAM dye, emission: 520 nm• Herpes simplex virus 2: yellow dye, emission: 549 nmTarget genes: • Glycoprotein B (UL 27, Herpes simplex virus 1)• Glycoprotein B (UL 27, Herpes simplex virus 2)Number of wells by test: 1 (Herpes simplex virus 1/ Herpes simplex virus 2)Samples used:• Bronchoalveolar lavages• Tissue/biopsy• Vesicular fluid• Ocular fluid• Swab corneal scraping• Cerebrospinal fluid• Amniotic fluid• Plasma• SerumValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® UltraSens Virus Kit (Qiagen)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm23

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011HERPES VIRUSESVaricella zoster virusRef: Dia-VZV-050Varicella zoster virus (VZV) is one of eight herpes viruses known toinfect humans. It commonly causes chicken-pox in children andboth shingles and postherpetic neuralgia in adults.Primary VZV infection results in chickenpox (varicella), which mayrarely result in complications including encephalitis or pneumonia.Even when clinical symptoms of chickenpox have resolved, VZVremains dormant in the nervous system of the infected person(virus latency), in the trigeminal and dorsal root ganglia (Steineret al., 2007). In about 10-20% of cases, VZV reactivates later in lifeproducing a disease known as herpes zoster or shingles. Seriouscomplications of shingles include postherpetic neuralgia, zostermultiplex, myelitis, herpes ophthalmicus, or zoster sine herpete.Steiner I, Kennedy PG, Pachner AR (2007). «The neurotropic herpes viruses:herpes simplex and varicella-zoster». Lancet Neurol 6 (11): 1015–28.VALIDATIONResearched pathogen:• Varicella zoster virus: FAM dye, emission: 520 nmTarget genes:• Glycoprotein B (ORF 31 gene, Varicella zoster virus)Number of wells by test: 1 (Varicella zoster virus)Samples used:• Bronchoalveolar lavage• Tissue/biopsy• Vesicular fluid• Ocular fluid• Swab corneal scraping• Cerebrospinal fluid• Amniotic fluid• Plasma• SerumValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® UltraSens Virus Kit (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm24 | www.diagenodediagnostics.com

BordetellaRef: Dia-Bor-020Whooping-cough is a highly contagious respiratory illness causedby the gram-negative bacterial pathogen Bordetella pertussis(Cherry, 2004). Among the nine species of Bordetella identifiedtoday, only three additional members (B.parapertussis, B.holmesii,B.bronchiseptica) have been associated with respiratory infectionsin humans and others mammals.Cherry, J. D., and U.Heininger. (2004). Pertussis and other Bordetella infections.In R.D.Feigin, J.D.Cherry, G.J.Demmler, and S.Kaplan (ed.), Textbook ofpediatric infectious diseases, 1588-1608.VALIDATIONResearched pathogens:Target genes:• Bordetella pertussis: FAM dye, emission: 520 nm• Bordetella parapertussis: FAM dye, emission: 520 nm• IS481 (Bordetella pertussis)• IS1001 (Bordetella parapertussis)Number of wells by test: 2 (Bordetella pertussis and Bordetella parapertussis)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 20 reactions (50 µl PCR, final volume)• 40 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmRESPIRATORY INFECTIONS25

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSBordetella multiplexRef: Dia-Bor-050Whooping-cough is a highly contagious respiratory illness causedby the gram-negative bacterial pathogen Bordetella pertussis(Cherry, 2004). Among the nine species of Bordetella identifiedtoday, only three additional members (B.parapertussis, B.holmesii,B.bronchiseptica) have been associated with respiratory infectionsin humans and others mammals.Cherry, J. D., and U.Heininger. (2004). Pertussis and other Bordetella infections.In R.D.Feigin, J.D.Cherry, G.J.Demmler, and S.Kaplan (ed.), Textbook ofpediatric infectious diseases, 1588-1608.VALIDATIONResearched pathogens:Target genes:• Bordetella pertussis: FAM dye, emission: 520 nm• Bordetella parapertussis: yellow dye, emission: 549 nm• IS481 (Bordetella pertussis)• IS1001 (Bordetella parapertussis)Number of wells by test: 1 (Bordetella pertussis and Bordetella parapertussis)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• Amplicor respiratory specimen preparation kit (RSP prep) (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm26 | www.diagenodediagnostics.com

Human influenza A/BRef: Dia-Inf.A/B-050Human influenza A/B (HI.A/B) is a respiratory infection of upperand lower respiratory tract in infants and young children. It causesbronchiolitis and pneumonia.Influenza is a negative-sense, enveloped RNA virus. Its morphologyis spherical or filamentous with a diameter of 80 to 120 nm. Itsmost severe symptoms are associated with influenza A viruses.Many sub-types of influenza A viruses are known and classifiedaccording to the origin of the hemagglutinin and neuraminidase.Influenza B viruses are antigenically more stable than influenzaA. This explains that they cause less severe epidemics.VALIDATIONResearched pathogens:• Human influenza A: FAM dye, emission: 520 nm• Human influenza B: yellow dye, emission: 549 nmTarget genes: • MP (Human influenza A)• NP (Human influenza B)Number of wells by test: 1 (Human influenza A / Human influenza B)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm27

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSHuman metapneumovirusRef: Dia-MPV-050Human metapneumovirus (MPV) is a respiratory infection ofupper and lower respiratory tract in infants and young children. Itcauses bronchiolitis and pneumonia.Human metapneumovirus is a negative-sense, single strandedRNA virus. Early assessments suggest that this virus maybe similar to respiratory syncytial virus in having two distinctgenotypes and a similar epidemiology.VALIDATIONResearched pathogen:• Human metapneumovirus: FAM dye, emission: 520 nmTarget genes:• N (Human metapneumovirus)Number of wells by test: 1 (Human metapneumovirus)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm28 | www.diagenodediagnostics.com

Human parainfluenza 1/3Ref: Dia-HPIV(1/3)-050Human parainfluenza 1/3 (HPIV (1/3)) is a respiratory infection ofupper and lower respiratory tract in infants and young children. Itcauses bronchiolitis and pneumonia.HPIVs are a group of four different serotypes of single-strandedRNA viruses belonging to the paramyxovirus family. They are thesecond most common cause of lower respiratory tract infectionin younger children.VALIDATIONResearched pathogens:• Human parainfluenza 1: FAM dye, emission: 520 nm• Human parainfluenza 3: yellow dye, emission: 549 nmTarget genes: • HN (Human parainfluenza 1)• HN (Human parainfluenza 3)Number of wells by test: 1 (Human parainfluenza 1/ Human parainfluenza 3)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm29

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSHuman parainfluenza 2/4Ref: Dia-HPIV(2/4)-050Human parainfluenza 2/4 (HPIV (2/4)) is a respiratory infection ofupper and lower respiratory tract in infants and young children. Itcauses bronchiolitis and pneumonia.HPIVs are a group of four different serotypes of single-strandedRNA viruses belonging to the paramyxovirus family. They are thesecond most common cause of lower respiratory tract infectionin younger children.VALIDATIONResearched pathogens:• Human parainfluenza 2: FAM dye, emission: 520 nm• Human parainfluenza 4: yellow dye, emission: 549 nmTarget genes: • HN (Human parainfluenza 2)• Phosphoprotein (Human parainfluenza 4)Number of wells by test: 1 (Human parainfluenza 2/ Human parainfluenza 4)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm30 | www.diagenodediagnostics.com

Human respiratory syncytialvirusRef: Dia-RSV-050Human respiratory syncytial virus (RSV) is a respiratory infectionof upper and lower respiratory tract in infants and young children.It causes bronchiolitis and pneumonia.Human respiratory syncytial virus is a negative-sense, envelopedRNA virus. The virion is variable in shape and size (averagediameter from 120 to 300 nm). It is also unstable in the environment(surviving only a few hours on environmental surfaces).VALIDATIONResearched pathogen:• Human respiratory syncytial virus: FAM dye, emission: 520 nmTarget genes:• N (Human respiratory syncytial virus)Number of wells by test: 1 (Human respiratory syncytial virus)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm31

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSHuman rhinovirusRef: Dia-HRV-050Human rhinovirus (HRV) is a respiratory infection of upper andlower respiratory tract in infants and young children. It causesbronchiolitis and pneumonia.HRV is a positive-sense, single strand RNA virus within anicosahedral (20-sided) capsid. Approximately 101 serotypes arecurrently identified.VALIDATIONResearched pathogen:• Human rhinovirus: FAM dye, emission: 520 nmTarget genes:• 5’UTR (Human rhinovirus)Number of wells by test: 1 (Human rhinovirus)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm32 | www.diagenodediagnostics.com

LegionellaRef: Dia-Lpn-050Legionella (Gram negative bacterium), including species thatcause legionellosis (most notably Legionella pneumophila) hasbeen shown to cause community-, travel-, and nosocomiallyacquired pneumoniae. Infection of humans is usually via inhalationof aerosols from a variety of natural or water systems (Templeton,Scheltinga et al. 2003).Templeton, K. E., S. A. Scheltinga, et al. (2003). “Development and clinicalevaluation of an internally controlled, single-tube multiplex real-time PCRassay for detection of Legionella pneumophila and other Legionella species.”J Clin Microbiol 41(9): 4016-21.VALIDATIONResearched pathogens:Target genes:• Legionella species: FAM dye, emission: 520 nm• Legionella pneumophila: yellow dye, emission: 549 nm• P1 (Legionella species)• 16S (Legionella pneumophila)Number of wells by test: 1 (Legionella species / Legionella pneumophila)Samples used:• Bronchoalveolar lavage• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm33

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSMycobacterium tuberculosisRef: Dia-MT-050Mycobacterium tuberculosis (MTB) is a pathogenic bacterialspecies of the genus Mycobacterium and the causative agent ofmost cases of tuberculosis.Tuberculosis is a common and severe infectious disease thatusually attacks the lungs but can also affect the central nervoussystem, the lymphatic system, the circulatory system, thegenitourinary system, the gastrointestinal system, bones, joints,and even the skin.Other mycobacteria such as Mycobacterium bovis, Mycobacteriumafricanum, Mycobacterium canetti, and Mycobacterium microtialso cause tuberculosis, but these species are less common.VALIDATIONResearched pathogen:• Mycobacterium tuberculosis: FAM dye, emission: 520 nmTarget genes:• IS6110 (Mycobacterium tuberculosis)Number of wells by test: 1 (Mycobacterium tuberculosis)Samples used:• Bronchoalveolar lavage• SputumValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm34 | www.diagenodediagnostics.com

Mycoplasma pneumoniae &Chlamydophila pneumoniaeRef: Dia-MCpn-050Mycoplasma pneumoniae and Chlamydophila pneumoniae arethe organisms responsible for most of the cases of atypicalpneumonia in childrenAtypical pneumonia due to these bacteria usually causes mildforms of pneumonia and are characterized by a more protractedcourse of symptoms unlike other forms of pneumonia which canprogress more quickly with more severe early symptoms.Mycoplasma pneumoniae can give rise to mild upper respiratorytract infection, bronchitis, bronchiolitis and bronchopneumonia.The disease usually starts as influenza like syndrome withfever, illness, headache, scratchy sore throat and cough as thepredominant symptoms.Chlamydophila pneumoniae has been associated with pharyngitis,sinusitis and bronchitis.VALIDATIONResearched pathogens:Target genes:• Mycoplasma pneumoniae: FAM dye, emission: 520 nm• Chlamydophila pneumoniae: yellow dye, emission: 549 nm• P1 (Mycoplasma pneumoniae)• 16S (Chlamydophila pneumoniae)Number of wells by test: 1 (Mycoplasma pneumoniae / Chlamydophila pneumoniae)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• Amplicor respiratory specimen preparation kit (RSP prep) (Roche)• MagNa pure LC system (Roche)• QiaSymphony (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm35

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011RESPIRATORY INFECTIONSRespiratory RNA virusesmultiplexRef: Dia-ResRNAVir-050Human influenza (HI) A/B, human respiratory syncytial virus(RSV), human metapneumovirus (MPV), human rhinovirus (HRV)and human parainfluenza (HPIV) 1/2/3/4 are respiratory infectionsof upper and lower respiratory tract in infants and young children.They cause bronchiolitis and pneumonia.Influenza is a negative-sense, enveloped RNA virus. Its morphologyis spherical or filamentous with a diameter of 80 to120 nm. Itsmost severe symptoms are associated with influenza A viruses.Many sub-types of influenza A viruses are known and classifiedaccording to the origin of the hemagglutinin and neuraminidase.Influenza B viruses are antigenically more stable than influenza A.This explains that they cause less severe epidemics.RSV is a negative-sense, enveloped RNA virus. The virion isvariable in shape and size (average diameter from 120 to 300 nm).It is also unstable in the environment (surviving only a few hourson environmental surfaces).MPV is a negative-sense, single stranded RNA virus. Earlyassessments suggest that this virus may be similar to respiratorysyncytial virus in having two distinct genotypes and a similarepidemiology.HRV is a positive-sense, single strand RNA virus within anicosahedral (20-sided) capsid. Approximately 101 serotypes arecurrently identified.HPIVs are a group of four different serotypes of single-strandedRNA viruses belonging to the paramyxovirus family. They are thesecond most common cause of lower respiratory tract infectionin younger children.36 | www.diagenodediagnostics.comVALIDATIONResearched pathogens:• Human influenza A: FAM dye, emission: 520 nm• Human influenza B: yellow dye, emission: 549 nm• Human respiratory syncytial virus: FAM dye, emission: 520 nm• Human metapneuvirus: FAM dye, emission: 520 nm• Human rhinovirus: yellow dye, emission: 549 nm• Human parainfluenza 1: FAM dye, emission: 520 nm• Human parainfluenza 2: FAM dye, emission: 520 nm• Human parainfluenza 3: yellow dye, emission: 549 nm• Human parainfluenza 4: yellow dye, emission: 549 nm(RNA extraction & PCR inhibition control: yellow dye, emission: 549 nm)Target genes: • MP (Human influenza A)• NP (Human influenza B)• N (Human respiratory syncytial virus)• N (Human metapneumovirus)• HN (Human parainfluenza 1)• HN (Human parainfluenza 2)• HN (Human parainfluenza 3)• Phosphoprotein (Human parainfluenza 4)Number of wells by test: 51. Human influenza A/ Human influenza B2. Human respiratory syncytial virus/ RNA extraction & PCR inhibition control3. Human metapneuvirus/ Human rhinovirus4. Human parainfluenza 1/ Human parainfluenza 35. Human parainfluenza 2/ Human parainfluenza 4Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® Viral RNA Mini Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)

Swine influenzaRef: Dia-swH1N1-050Swine influenza (also called H1N1 flu, swine flu) is an infection byany one of several types of swine influenza virus. Swine influenzavirus (SIV) is any strain of the influenza family of viruses thatis endemic in pigs. As of 2009, the known SIV strains includeinfluenza C and the subtypes of influenza A known as H1N1,H1N2, H3N1, H3N2, and H2N3 (1) .Transmission of the virus from pigs to humans is not commonand does not always lead to human influenza, often resulting onlyin the production of antibodies in the blood. If transmission doescause human influenza, it is called zoonotic swine flu.Swine influenza. The Merck Veterinary Manual. 2008. Retrieved on April 30, 2009.http://www.merckvetmanual.com/mvm/index.jsp?cfile=htm/bc/121407.htm.VALIDATIONResearched pathogen:• Swine H1N1 influenza A virus: FAM dye, emission: 520 nmTarget genes:• HA (Swine H1N1 influenza A virus)Number of wells by test: 1 (Swine H1N1 influenza A virus)Samples used:• Bronchoalveolar lavage• Nasopharyngeal swabs/washesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® Viral RNA Mini Kit (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm37

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011MENINGITISNeisseria meningitidis /Serogroup B & CRef: Dia-Men-020Neisseria meningitidis (gram-negative bacteria) may leadto meningitis, and is responsible for considerable mortalitythroughout the world. Most disease-causing Neisseirameningitidis strains belong to serogroups B and C in Europe,North & South America and Australia.VALIDATIONResearched pathogens:Target genes:• Neisseria meningitidis (all strains): FAM dye, emission: 520 nm• Serogroup B: FAM dye, emission: 520 nm• Serogroup C: FAM dye, emission: 520 nm• CtrA (Neisseria meningitidis)• siaD (Serogroup B)• siaD (Serogroup C)Number of wells by test: 3 (Neisseria meningitidis and serogroup B and serogroup C)Samples used:• Human cerebrospinal fluid• Whole bloodValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 20 reactions (50 µl PCR, final volume)• 40 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm38 | www.diagenodediagnostics.com

Neisseria meningitidis /Serogroup A, B, C, W135 & YRef: Dia-MenSerogroup-020Neisseria meningitidis (gram-negative bacteria) may leadto meningitis, and is responsible for considerable mortalitythroughout the world. Most disease-causing Neisseirameningitidis strains belong to serogroups B and C in Europe,North & South America and Australia.VALIDATIONResearched pathogens:Target genes:• Neisseria meningitidis (all strains): FAM dye, emission: 520 nm• Serogroup A: FAM dye, emission: 520 nm• Serogroup B: FAM dye, emission: 520 nm• Serogroup C: FAM dye, emission: 520 nm• Serogroup W135: FAM dye, emission: 520 nm• Serogroup Y: FAM dye, emission: 520 nm• CtrA (Neisseria meningitidis)• sacB (Serogroup A)• siaD (Serogroup B)• siaD (Serogroup C)• synG (Serogroup W135)• synF (Serogroup Y)Number of wells by test: 6 (Neisseria meningitidis and serogroup A, B, C, W135 and Y)Samples used:• Human cerebrospinal fluid• Whole bloodValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 20 reactions (50 µl PCR, final volume)• 40 reactions (25 µl PCR, final volume)• 50 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm39

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011MENINGITISNeisseria meningitidis &Streptococcus pneumoniaeRef: Dia-MenMultiplex-050Neisseria meningitidis (gram-negative bacteria) may leadto meningitis, and is responsible for considerable mortalitythroughout the world. Most disease-causing Neisseirameningitidis strains belong to serogroups B and C in Europe,North & South America, and Australia.Streptococcus pneumoniae (Gram-positive bacteria) wasrecognized as a major cause of pneumonia in the late 19th centuryand is the most common cause of bacterial meningitis in adultsand children.VALIDATIONResearched pathogens:Target genes:• Neisseria meningitidis: FAM dye, emission: 520 nm• Streptococcus pneumoniae: yellow dye, emission: 549 nm• CtrA (Neisseria meningitidis)• Ply (Streptococcus pneumoniae)Number of wells by test: 1 (Neisseria meningitidis / Streptococcus pneumoniae)Samples used:• Human cerebrospinal fluid• Whole bloodValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nm40 | www.diagenodediagnostics.com

AdenovirusRef: Dia-ADV-050Human adenoviruses cause a variety of diseases and are prevalentthroughout the world.These non-enveloped and double-stranded DNA viruses arepathogens involved in conjunctivitis, keratoconjunctivitis,gastroenteritis, respiratory tract infections.Human adenoviruses are also associated with pneumonia,hemorrhagic cystitis, hepatitis, immunodeficienty and bonemarrow and solid organ transplant. (Claas et al., 2005; Sarantiset al., 2004).Claas, E. C., Schilham, M. W., de Brouwer, C. S., Hubacek, P., Echavarria, M.,Lankester, A. C., van Tol, M. J. and Kroes, A. C. (2005). Internally controlledReal-time PCR monitoring of adenovirus DNA load in serum or plasma oftransplant recipients. J Clin Microbiol 43, 1738-44.Sarantis, H., Johnson, G., Brown, M., Petric, M. and Tellier, R. (2004).Comprehensive detection and serotyping of human adenoviruses by PCR andsequencing. J Clin Microbiol 42, 3963-9.VALIDATIONResearched pathogen:• Adenovirus: FAM dye, emission: 520 nmTarget genes:• Exon & VA RNA (Adenovirus)Number of wells by test: 1 (Adenovirus)Samples used:• Bronchoalveolar lavage• Sputum• Nasopharyngeal swabs/washes• Whole blood• Plasma• UrinesValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® UltraSens Virus Kit (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (2.0/480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(TR)-050: texas red dye, emission: 603 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(TR)-050: texas red dye, emission: 603 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmOTHER PATHOGENS41

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011OTHER PATHOGENSBorrelia burgdorferiRef: Dia-BorBurg-050Borrelia burgdorferi is a species of Gram negative bacteria of thespirochete class of the genus Borrelia.B. burgdorfer is predominant in North America, but also exists inEurope, and is the agent of Lyme disease.VALIDATIONResearched pathogens:• Borrelia burgdorferi: FAM dye, emission: 520 nmTarget genes:• ospA (Borrelia burgdorferi)Number of wells by test: 1 (Borrelia burgdorferi)Samples used:• cerebrospinal fluidValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® DNA Mini Kit (50) (Blood and Body Fluid Spin Protocol) (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmDIA-EIC/DNA(RD)-050: red dye, emission: 669 nm42 | www.diagenodediagnostics.com

EnterovirusRef: Dia-ENT-050The enteroviruses are associated with several human diseases.Historically the most significant has been the poliovirus.Enteroviruses are the most common cause of aseptic meningitisand can cause serious disease especially in infants.Serologic studies have distinguished 66 human enterovirusserotypes on the basis of an antibody neutralization test, andadditional antigenic variants have been defined within severalof the serotypes on the basis of reduced or nonreciprocal crossneutralizationbetween prototype and variant strains.Enteroviruses can be found in the respiratory secretions (saliva,sputum, nasal mucus) and stool- of an infected person (1)(2) .(1)Oberste MS, Maher K, Kilpatrick DR, Pallansch MA (1999). “Molecularevolution of the human enteroviruses: correlation of serotype with VP1sequence and application to picornavirus classification”. J. Virol. 73 (3): 1941–8.(2)CDC article Non-Polio Enterovirus Infections.VALIDATIONResearched pathogen:• Enterovirus: FAM dye, emission: 520 nmTarget genes:• 5’UTR (Enterovirus)Number of wells by test: 1 (Enterovirus)Samples used:• Tissue• Vesicular fluid• Cerebrospinal fluid• Swab• Feces• Plasma• SerumValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• MagNa pure LC system (Roche)• QIAamp ® Viral RNA Mini Kit (Qiagen)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)PCR volumes: 50 µl, 25 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• RNA extraction & PCR inhibition control (DIA-EIC/RNA)-050:DIA-EIC/RNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/RNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/RNA(Cy5)-050: Cy5 dye, emission: 662 nm43

HUMAN MOLECULAR DIAGNOSTICS CATALOGUE 2011OTHER PATHOGENSParvovirus B19Ref: Dia-PVB19-050The B19 virus, generally referred to as parvovirus B19 (1) orsometimes erythrovirus B19 (2) , was the first (and until 2005 theonly) known human virus in the family of parvoviruses, genuserythrovirus. B19 virus causes a childhood rash called fifthdisease or erythema infectiosum which is commonly calledslapped cheek syndrome (3) .(1)Servey JT, Reamy BV, Hodge J (February 2007). “Clinical presentations ofparvovirus B19 infection”. Am Fam Physician 75 (3): 373–6. (2) Kahn JS, KesebirD, Cotmore SF, et al. (July 2008). “Seroepidemiology of human bocavirusdefined using recombinant virus-like particles”. J. Infect. Dis. 198 (1): 41–50.(3)Vafaie J, Schwartz RA (2004). “Parvovirus B19 infections”. Int J Dermatol 43(10): 747–9.VALIDATIONResearched pathogens:• Parvovirus B19: FAM dye, emission: 520 nmTarget genes: • NS (Parvovirus B19)Number of wells by test: 1 (Parvovirus B19)Samples used:• EDTA plasma• SerumValidated extraction methods: • Nuclisens easyMAG ® System (Biomerieux)• QIAamp ® UltraSens Virus Kit (Qiagen)• MagNa pure LC system (Roche)Real-time PCR systems: • ABI (7000/7300/7500/7900)• Roche LightCycler ® (480/2.0 on request)• Biorad (iCycler/IQ5/CFX96)• Qiagen Rotor-Gene (6000)• Stratagene (MX3000P/3005P)• Cepheid (SmartCycler II)PCR volumes: 50 µl, 25 µl, 20 µl (depending of the real-time PCR system used)PCR reactions/kit:• 50 reactions (50 µl PCR, final volume)• 100 reactions (25 µl PCR, final volume)• 120 reactions (20 µl PCR, final volume)In combination with:• Universal inhibition control (DIA-UIC-050):DIA-UIC(YD)-050: yellow dye, emission: 549 nmDIA-UIC(DR)-050: orange dye, emission: 576 nmDIA-UIC(Cy5)-050: Cy5 dye, emission: 662 nm• DNA extraction & PCR inhibition control (DIA-EIC/DNA)-050:DIA-EIC/DNA(YD)-050: yellow dye, emission: 549 nmDIA-EIC/DNA(DR)-050: orange dye, emission: 576 nmDIA-EIC/DNA(Cy5)-050: Cy5 dye, emission: 662 nmDIA-EIC/DNA(RD)-050: red dye, emission: 669 nm44 | www.diagenodediagnostics.com

Diagenode’s wide range of kits enables new levels of throughput, cost savingsand ease-of-use for real-time qPCR IVD.HIGHER THROUGHPUT LOWER COSTALL EXTRACTION METHODS ALL qPCR PLATFORMSALL MASTER MIXEST0T130’ 60’QC in the Extraction Step*Patient sampleDiagenode’s extractionand inhibition controlSmart qPCR Set-UpOur kits are designed and validated for a variety ofreal-time qPCR platforms** as well as for differentextraction protocols. The kits are validated with themost commonly-used master mixes. To achieve optimalsensitivity with our kits, we provide recommendations onwhich master mixes will provide the best results.KIT 3Ex: LegionellaKIT 1Ex: Varicella zoster virusKIT 2Ex: CytomegalovirusOnly one set of qPCR cycles for ALL Diagenode’s kits allows costand time savings. The same PCR plate can be used for testingdifferent pathogens.SampleBPositive controlDiagenode suggests using DNA or RNA extraction andinhibition controls (Dia-EIC/DNA-050, Dia-EIC/RNA-050)for highest test accuracy. These controls are availablewith different dyes: YD, Cy5, DR and TR (see page 31).Note: If you do not use an extraction control, you can useour universal inhibition control (Dia-UIC-050). This contolis available with different dyes: YD, Cy5, DR and TR.Human sampleDiagenode’s primers andprobes (pathogen + ctl***)Master mix(Your favorite**)Diagenode’s positivecontrolDiagenode’s primers andprobes (pathogen + ctl***)Master mix(Your favorite**)* Most standard extraction methods are compatible withDiagenode’s kits.** Most have been validated with our kits. Ask for results!*** ctl = Dia-EIC/DNA-050, Dia-EIC/RNA-050 or Dia-UIC-050

Diagenode’s expertise the secure way to performanceHIGH SENSITIVITYFOR MONO OR MULTIPLEX qPCRACCURATE FAST CONVENIENTT2150’T3< 3hEasy qPCR curve analysisFast results for clinicians and patients0.8High precision detection of Human Cytomegalovirususing kit # dia-CMVQ-050In less than 3 hours, including extraction step, physicians can decide the mostappropriate therapeutic strategy for his patient.Norm. Fluoro0.60.4The figure shows detection of 5 dilutions (induplicate) of the positive control (standard curve)processed with 1) eight samples containingvarious concentrations of CMV and 2) twonegative samples from the EuropeanQuality Control for Molecular Diagnostics(QCMD)*Customerfeedback“I love the flexibility of the Diagenode IVD kits. Our lab uses differentqPCR machines and two different extraction robots, yet Diagenode’skits perform with high consistency each and every time across thedifferent machines.0.20.0* Including the extraction stepQCMDStandard curve Dia-CMVQ-050Threshold510 15 20 25Cycle30 35 40 45At first, I was not sure whether Diagenode would be able to competewith the larger IVD companies. We quickly realized that the Diagenodekits deliver excellent performance with consistent and absolutelyreliable results, for a fraction of the cost of competing kits! Diagenodeexceeded our expectations with kits that performed better orcomparably to those of the larger suppliers.CT353025Standard Curve (2) CT = -3.256*log(conc) + 37.351Reaction efficiency* 1.02835 (* = 10^(-1/m) - 1)M -3.25581B 37.35086R Value 0.99624R^2 Value 0.99249In addition, Diagenode has a long history of expertise in nucleic acidtesting, adding technological power to their IVD kits and assuring thehighest level of test accuracy. The Diagenode in-house experts provideuseful experimental tips and guidance in adapting their products forour specific technological needs.”2010 1 10 2 10 3 10 410 5 10 6Diagenode customerStandard curveConcentration

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