Package insert

Nanogen Advanced Diagnostics S.r.L. 

Corso Torino, 89/d 

10090 Buttigliera Alta (TO) ITALY 

Offices: 

Tel. +39-011 976 19 1 

Fax +39-011 936 76 11 

E. mail: techsupport@nanogenad.com 

Q - PCR Alert AmpliMASTER 

reagents optimized for real time amplification 

Complete size product, code RTS000 

MATERIALS PROVIDED IN THE PRODUCT 

RTS000 

RTS000/2 

«REAL TIME AMPLIFICATION ACCESSORIES» 



RTS000 

RTS000/2 

web site: www.nanogen.com 

TABLE OF CONTENTS 

INTENDED USE page 1 

PRODUCT DESCRIPTION page 1 

MATERIALS PROVIDED IN THE PRODUCT page 2 

MATERIALS REQUIRED BUT NOT PROVIDED IN THE PRODUCT page 2 

ACCESSORY PRODUCTS page 2 

WARNINGS AND PRECAUTIONS page 3 

PROCEDURE page 4 

SYMBOLS page 4 

INTENDED USE 

«Q - PCR Alert AmpliMASTER» is part of the quantitative amplification assay of nucleic acids for 

the detection and dosing of a target DNA with the real time amplification method in samples of DNA 

extract or cDNA obtained from RNA extract and for allele determination with real time amplification 

method in samples of DNA extract. The product is an accessory of the products in the «Q - PCR Alert 

AmpliMIX», «Q - PCR Alert AmpliPROBE», «Q - PCR Standard» lines and in the «Positive Control» lines 

by Nanogen Advanced Diagnostics S.r.L. 

The product is intended for use in the preparation of diagnostic assays based on the real time 

amplification method. 

PRODUCT DESCRIPTION 

+2° C 

+8° C 

In the complete size (RTS000) the product provides the mixture of reagents optimized for 

AmpliMASTER real time amplification in a stabilizing solution divided into aliquots in four ready-to-use test 

tubes, three amplification microplates with 96 wells and three adhesive sheets for amplification. 

In the reduced size (RTS000/2) the product provides the mixture of reagents optimized for 

AmpliMASTER real time amplification in a stabilizing solution divided into aliquots in two ready-to-use test 

tubes, two amplification microplates with 96 wells and two adhesive sheets for amplification. 

The reagent mixture provides the buffer system, magnesium chloride, triphosphate nucleotides, ROX 

fluorophore as the passive reference for the normalisation of the fluorescence, the enzyme Uracil N- 

glycosidase (UNG) to inactivate contamination from the amplification product, the enzyme Taq DNA 

polymerase for hot start. 

In the complete size (RTS000) the product provides 96 determinations, including standards and 

controls. 

In the reduced size (RTS000/2) the product provides 48 determinations, including standards and 

controls. 

SCH mRTS000_en 20/04/07 Review 01 Page 1/4 

Component Description Quantity Composition Labelling 

AmpliMASTER optimized reagent mixture 4 x 340 µl 

Microplate for amplification 

microplate 

with 96 x 0.2 ml wells 

TRIS base, TRIS hydrochloride, 

Glycerol, magnesium chloride, 

Deoxyribonucleotide triphosphates, 

ROX, Uracil-N-glycosidase, Taq 

DNA polymerase 

3 - - 

Adhesive sheet for amplification adhesive sealing sheet 3 - - 

Reduced size product, code RTS000/2 


AmpliMASTER optimized reagent mixture 2 x 340 µl 

Microplate for amplification 

microplate 

with 96 x 0.2 ml wells 


Glycerol, magnesium chloride, 

Deoxyribonucleotide triphosphates, 

ROX, Uracil-N-glycosidase, Taq 

DNA polymerase 

2 - - 

Adhesive sheet for amplification adhesive sealing sheet 2 - - 

MATERIALS REQUIRED BUT NOT PROVIDED IN THE PRODUCT 

- Laminar airflow hood. 

- Disposable latex powder-free gloves or similar material. 

- Vortex mixer 

- Bench microcentrifuge (12,000 - 14,000 RPM). 

- Sterile micropipettes and tips with aerosol filter or positive displacement (0.5-10 µl, 2-20 µl, 5-50 µl, 

50-200 µl, 200-1000 µl). 

- Sterile bidistilled water. 

- Programmable heater with optical fluorescence detection system (thermal cycler for real time). 

ACCESSORY PRODUCTS 

The primer reagents (oligonucleotides), the detection reagents (fluorescent probes), the knownquantity 

DNA standard and the positive controls on the plasmid DNA, are not included in this product. To 

perform these analytical steps the following accessory products, manufactured by Nanogen Advanced 

Diagnostics S.r.L., are recommended: 

«Q - PCR Alert AmpliMIX» series, primer oligonucleotides for real time amplification. 

«Q - PCR Alert AmpliPROBE» series, fluorescent probes for real time amplification. 

«Q - PCR Standard» series, known-quantity plasmid DNA to obtain the standard curve. 

«Positive Control» series, positive control of plasmid DNA. 


- 

-



RTS000 

RTS000/2 



RTS000 

RTS000/2 

This product is exclusively for in vitro use. 

Warnings and general precautions 

WARNINGS AND PRECAUTIONS 

Handle and dispose of all biological samples as if they were capable of transmitting infective agents. 

Avoid direct contact with the biological samples. Avoid splashing or spraying. The materials that come into 

contact with biological samples must be treated with 3% sodium hypochlorite for at least 30 minutes or 

autoclaved at 121°C for one hour before disposal. 

Handle and dispose of all reagents and all assay materials as if they were capable of transmitting 

infective agents. Avoid direct contact with the reagents. Avoid splashing or spraying. Waste must be treated 

and disposed of in compliance with the appropriate safety standards. Disposable combustible materials must 

be incinerated. Liquid waste containing acids or bases must be neutralised before disposal. 

Wear suitable protective clothing and gloves and protect eyes / face. 

Never pipette solutions by mouth. 

Do not eat, drink, smoke or apply cosmetic products in the work areas. 

Wash hands carefully after handling samples and reagents. 

Dispose of leftover reagents and waste in compliance with regulations in force. 

Read all the instructions provided with the product before running the assay. 

Follow the instructions provided with the product while running the assay. 

Do not use the product after the expiry date. 

Only use the reagents provided in the product and those recommended by the manufacturer. 

Do not mix reagents from different batches. 

Do not use reagents from other manufacturers' products. 

Warnings and precautions for molecular biology 

Molecular biology procedures, such as extraction, reverse transcription, amplification and detection 

of nucleic acids, require qualified staff to prevent the risk of erroneous results, especially due to degradation 

of the nucleic acids contained in the samples or due to sample contamination by amplification products. 

It is necessary to have separate areas for the extraction/preparation of amplification reactions and for 

the amplification/detection of amplification products. Never introduce an amplification product in the area 

designed for extraction/preparation of amplification reactions. 

It is necessary to have lab coats, gloves and tools which are exclusively employed in the 

extraction/preparation of amplification reactions and for the amplification/detection of amplification products. 

Never transfer lab coats, gloves or tools from the area designed for the amplification/detection of 

amplification products to the area designed for the extraction/preparation of the amplification reactions. 

The samples must be exclusively employed for this type of analysis. Samples must be handled 

under a laminar flow hood. Tubes containing different samples must never be opened at the same time. 

Pipettes used to handle samples must be exclusively employed for this specific purpose. The pipettes must 

be of the positive displacement type or be used with aerosol filter tips. The tips employed must be sterile, 

free from DNases and RNases, free from DNA and RNA. 

Reagents must be handled under a laminar flow hood. The reagents required for amplification must 

be prepared in such a way that they can be used in a single session. The pipettes employed to handle the 

reagents must be used exclusively for this purpose. The pipettes must be of the positive displacement type 

or be used with aerosol filter tips. The tips employed must be sterile, free from DNases and RNases, free 

from DNA and RNA. 

Amplification products must be handled in such a way as to reduce dispersion into the environment 

as much as possible, in order to avoid the possibility of contamination. Pipettes used to handle amplification 

products must be employed exclusively for this specific purpose. 

Warnings and precautions specific to components 

AmpliMASTER does not carry risk phrases (R) and it carries the following safety warnings (S): 

S 23-25 Do not breathe gas/fumes/vapour/spray. Avoid contact with eyes. 

PROCEDURE 

The «Q - PCR Alert AmpliMASTER» product must be used with the products in the «Q - PCR Alert 

AmpliMIX» series and the «Q - PCR Alert AmpliPROBE» line to obtain the reaction mixture. 

AmpliMASTER is ready for use, hence must be used directly in the preparation of the reaction 

mixture. 

The complete procedure involves preparation and execution of a real time amplification reaction on a 

microplate with programmable heater with optical fluorescence detection system (thermal cycler for real time) 

and is described in detail in the instruction manual enclosed with the products in the «Q - PCR Alert 

AmpliMIX» series. 

The performance characteristics and procedure limitations of the assay including detection and 

dosing of the target DNA are described in detail in the instruction manual enclosed with the products in the 

«Q - PCR Alert AmpliMIX» series. 

Catalogue number. 

Temperature limits. 

Batch code. 

Use by (last day of month). 

In vitro diagnostic medical device. 

SYMBOLS 

In keeping with the requirements of European Directive 98\79\EC for in vitro diagnostic 

medical devices. 

Contents sufficient for "N" tests. 

Please refer to the instructions for use. 

Manufacturer. 

The purchase of this product allows the purchaser to use it for amplification and detection of nucleic acid sequences providing human in 

vitro diagnostic services. This right is granted only if this product is used in association with Nanogen Advanced Diagnostics S.r.L. 

licensed products for "Positive Control" or “Q - PCR Standard”. 

No general patent or other license of any kind other then this specific right of use from purchase is granted hereby. 


SCH mRTS000_en 20/04/07 Review 01 Page 4/4




VZV Q - PCR Alert AmpliMIX 

detection and dosing of VZV DNA 

RTS035-M 

RTS035-M 

Offices: 

Tel. +39-011 976 19 1 

Fax +39-011 936 76 11 

E-mail: techsupport@nanogenad.com 


«DUPLEX REAL-TIME AMPLIFICATION» 



-20°C 

ASSAY PRINCIPLE 

The procedure involves a real-time amplification reaction on a microplate with programmable heater 

with optical fluorescence detection system (thermal cycler for real time). 

In each well, an amplification reaction is carried out specific for a gene region that codifies the Major 

DNA binding protein (ORF 29) of VZV and for a region of the human beta globin gene (internal suitability 

test of the sample) using the DNA extracted from the samples being tested. A specific probe for VZV labelled 

with FAM fluorophore is activated when hybridized with the specific product of the VZV amplification reaction. 

Another probe specific for the human gene of beta globin labelled with VIC fluorophore is activated when 

hybridized with the product of the amplification reaction for the human beta globin gene. Fluorescence 

emission increases as the specific products of the amplification reaction increase and is measured and 

recorded by the instrument. The processing of the data determines the presence and the titre of VZV DNA in 

the starting sample. 

System standardization was carried out on Applied Biosystems ABI PRISM TM 7000 series 

instruments. 




ASSAY PRINCIPLE page 2 






SAMPLES AND CONTROLS page 5 


PROCEDURE LIMITATIONS page 12 

PERFORMANCE CHARACTERISTICS page 13 

REFERENCES page 16 

TROUBLESHOOTING page 17 


INTENDED USE 

«VZV Q - PCR Alert AmpliMIX» is part of a quantitative amplification assay of nucleic acids for the 

detection and dosing of the DNA of human herpetic Varicella - Zoster virus (VZV) in DNA samples 

extracted from swabs of mucocutaneous lesions, cephalo-rachidian liquid and from plasma collected in 

EDTA. 

The product is intended for use, alongside clinical data and other laboratory tests, in the diagnosis 

and monitoring of VZV infections. 

The product supplies the mixture of AmpliMIX primer oligonucleotides for real time amplification in a 

stabilizing solution, pre-dosed in aliquots into four disposable test tubes. Each test tube contains 110 µl 

of solution, sufficient for 24 tests. 

The primer oligonucleotides for VZV are specific for a gene region that codifies the Major DNA 

binding protein (ORF29) of VZV. 

The primer oligonucleotides for the internal suitability test of the sample are specific for the promoter 

region and 5' UTR of the human beta globin gene. 

The product enables 96 determinations, including standards and controls. 



VZV AmpliMIX 

primer oligonucleotides 

mixture 

4 x 110 µl 

Oligonucleotides, 


Glycerol, Triton X-100 




- Vortex mixer. 

- Bench microcentrifuge (12,000 - 14,000 RPM). 


50-200 µl, 200-1000 µl). 


- Programmable heater with optical fluorescence detection system. 

- 

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SCH mRTS035M_en 11/12/06 Review 00 Page 2/18



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The reagents for DNA extraction from the samples to be analysed, the positive extraction control, the 

reagents optimized for amplification, the detection reagents (fluorescent probes) and the positive 

amplification control or the known-quantity DNA standard are not included in this product. To perform these 

analytical steps the following accessory products, manufactured by Nanogen Advanced Diagnostics S.r.L., 

are recommended: 

«EXTRAgen®» (code EXTG01), kit for extraction of nucleic acids from non-cellular samples; the kit 

enables 50 extractions. 

«CPE-DNA ® - Internal Control» (code CTREXTG), positive plasmid DNA extraction control for 

non-cellular sample DNA extractions; the product enables 50 extractions. 

«Q - PCR Alert AmpliMASTER» (code RTS000), combination of optimized reagents, microplates 

and adhesive sheets for real time amplification; the product enables 96 reactions. 

«VZV Q - PCR Alert AmpliPROBE» (code RTS035-P), primer oligonucleotides for real time 

amplification; the product enables 96 reactions. 

If a qualitative result of the analysis is required (detection of VZV DNA): 

«VZV - Positive Control» (code CTR035), positive amplification control of plasmid DNA; the product 

enables 25 sessions. 

If a quantitative result of the analysis is required (dosing of VZV DNA): 

«VZV Q - PCR Standard» (code STD035), known-quantity plasmid DNA to obtain the standard 

curve; the product enables 16 sessions. 








Handle and dispose of all reagents and all assay materials as if they were potentially infective. Avoid 

direct contact with the reagents. Avoid splashing or spraying. Waste must be treated and disposed of in 

compliance with the appropriate safety standards. Disposable combustible materials must be incinerated. 

Liquid waste containing acids or bases must be neutralised before disposal. 




































Warnings and precautions specific to components 

The test tubes containing AmpliMIX are disposable and therefore must be used once only in the 

preparation of the reaction mixture. 

AmpliMIX carries the following safety warnings (S): 






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SAMPLES AND CONTROLS 

PROCEDURE 

Samples 

This product must be used with DNA extracted from the following biological samples: swabs of 

mucocutaneous lesions, cephalo-rachidian liquid, plasma collected in EDTA. 

Swabs of mucocutaneous lesions 

The swabs of mucocutaneous lesions that are to be used for DNA extraction must be taken 

according to laboratory guidelines, resuspended in transport medium for cell cultures or sterile physiological 

solution or sterile PBS, transported at +2° / +8° C and stored at +2° / +8° C for a maximum of four ho urs 

otherwise they must be frozen and stored at -20° C for a maximum of thirty days or at -70° C for longe r 

periods. 

It is advisable to split the samples that are to be stored frozen into aliquots in order to prevent 

repeated cycles of freezing and thawing. 

Instructions for pre-treatment of clinical samples, where applicable, and for DNA extraction are 

contained in the instruction manual for «EXTRAgen®» kit. 

Cephalo-rachidian liquid 

The cephalo-rachidian liquid samples that are to be used for DNA extraction must be collected 

according to laboratory guidelines, avoiding contamination from the patient's blood, transported at +2° / +8° 

C and stored at +2° / +8° C for a maximum of four h ours otherwise they must be frozen and stored at -20° C 

for a maximum of thirty days or at -70° C for longe r periods. 



Cephalo-rachidian liquid samples do not require pre-treatment and may be used directly for DNA 

extraction according to the instructions contained in the manual for «EXTRAgen ® » kit. 

Plasma collected in EDTA 

The plasma collected in EDTA must be stored according to laboratory guidelines, transported at 

+2° / +8° C and stored at +2° / +8° C for a maximum of four hours or frozen at -20° C for a maximum of thirty 

days or at -70° C for longer periods. 



The plasma collected in EDTA do not require pre-treatment and may be used directly for DNA 

extraction according to the instructions contained in the manual for «EXTRAgen ® » kit. 

Interfering substances 

The DNA extracted from the starting sample must not contain heparin or haemoglobin in order to 

prevent the problem of inhibition and the possibility of frequent invalid results. 

There are no data available concerning inhibition caused by antiviral drugs, antibiotics, 

chemotherapeutic drugs or immunosuppressants. 

Amplification controls 

It is absolutely mandatory to validate each amplification session with a positive control reaction and a 

negative control reaction. 

For the negative control, use sterile bidistilled water (not supplied with product) added to the reaction 

in place of the DNA extracted from the sample. 

For the positive control, use the «VZV - Positive Control» product or the «VZV Q - PCR Standard» 

product. 

Quality controls 

It is recommended to validate the whole analysis procedure of each extraction and amplification 

session by processing a negative sample and a positive sample that have already been tested or calibrated 

reference material. 

Setting up the real time amplification session 

(To be performed in the amplification / detection area of the amplification products) 

Before starting the session it is important to do the following: 

- referring to the instrument documentation, switch on the real time thermal cycler, switch on the 

control computer, launch the software and open an "absolute quantification" session; 

- referring to the instrument documentation, set the "detector" for the VZV probe with the reporter as 

"FAM" and the "quencher" as "none" (NFQ = non-fluorescent quencher); 

- referring to the instrument documentation, set the “detector” for the beta globin probe with the 

“reporter” as "VIC" and the “quencher” as "none" (NFQ = non-fluorescent quencher); 

- referring to the instrument documentation, for each well used in the microplate, set the “detectors” 

(type of fluorescence that is to be measured), the “passive reference” as “ROX” (normalisation of 

measured fluorescence) and the type of reaction (sample, negative amplification control, positive 

amplification control, known-quantity standard). Add this information to the Work Sheet enclosed at 

the end of this instruction manual or print the microplate organisation. The Work Sheet must be 

followed carefully during the transfer of the reaction mixture and samples into the wells. 

N.B.: In order to determine the DNA titre in the starting sample, set up a series of reactions with the Q - PCR 

standards (10 5 copies, 10 4 copies, 10 3 copies, 10 2 copies) to obtain the standard curve. 

Below is an example of how the analysis of 11 samples can be organized. 

S1 S9 

S2 S10 

S3 S11 

S4 NC 

S5 10 2 

S6 10 3 

S7 10 4 

S8 10 5 

Key: S1 - S11: Samples to be analysed; NC: Negative amplification control; 

10 2 : Standard 10 2 copies; 10 3 : Standard 10 3 copies; 10 4 : Standard 10 4 copies; 10 5 : Standard 10 5 copies. 

- Referring to the instrument documentation, set the parameters of the thermal cycle on the thermalcycler 

and a reaction volume of 25 µl. For Applied Biosystems ABI PRISM TM instruments of the 7000 

series choose the "9600 emulation" option. 

Amplification thermal cycle 

Phase Temperature Timing 

Decontamination 50° C 2 min. 

Initial denaturation 95° C 10 min. 

45 cycles 

95° C 15 sec. 

60° C 1 min. 





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Amplification set-up 

(To be performed in the extraction/preparation area of the amplification reaction) 

Before starting the session it is important to do the following: 

- remove and thaw the test tubes containing the samples to be analysed. Centrifuge the tubes to 

bring the contents to the bottom and keep in ice; 

- remove and thaw the AmpliMIX test tubes needed for the session, remembering that the contents 

of each tube are sufficient for 24 reactions. Centrifuge the tubes for 5 seconds to bring the contents 

to the bottom and keep in ice; 

- remove and thaw the same number of test tubes of AmpliPROBE as the AmpliMIX tubes. 

Centrifuge the tubes for 5 seconds to bring the contents to the bottom and keep in ice; 

- remove the same number of test tubes of AmpliMASTER as the AmpliMIX tubes. Write "VZV" 

and the date on the test tube label using indelible ink. Centrifuge the tubes for 5 seconds to bring the 

contents to the bottom and keep in ice; 

- remove and thaw the Positive Control test tube or the Q - PCR Standard tubes. Centrifuge the 

tubes for 5 seconds to bring the contents to the bottom and keep in ice; 

- If necessary, cut the amplification microplate to separate the part that will be used in the session, 

being careful to handle it with powder-free gloves and not to damage the wells. 

1. Transfer 100 µl of AmpliMIX to the AmpliMASTER tube. Mix well and pipette the volume of 100 µl 

three times into the mix. 

2. Transfer 100 µl of AmpliPROBE to the AmpliMASTER tube. Mix well and pipette the volume of 100 µl 

three times into the mix. 

3. Vortex on a low setting for 5 seconds, avoiding the creation of foam. 

4. Centrifuge the tubes for 5 seconds to bring the contents to the bottom. 

5. Gently deposit 20 µl of the reaction mixture obtained in this way on the bottom of the amplification 

microplate wells, as previously established on the Work Sheet. 

N.B.: If not all the reaction mixture is used, store the remaining volume in the dark at -20° C for a maxim um 

of one month in the test tube labelled "VZV". Freeze and thaw the reaction mixture ONLY ONCE. 

6. Gently deposit 5 µl of DNA extract from the first sample in the reaction mixture in the corresponding 

well of the amplification microplate, as previously established on the Work Sheet. Proceed in this 

way for all the other DNA extracts. 

7. Gently deposit 5 µl of sterile bidistilled water (not supplied with the product) in the reaction mixture in 

the well of the negative control amplification microplate, as previously established on the Work 

Sheet. 

8. Gently deposit 5 µl of Positive Control in the reaction mixture in the corresponding amplification 

microplate well, as previously established on the Work Sheet. 

N.B.: When this product is used for dosing VZV DNA, carefully deposit 5 µl of Q - PCR Standard 10 2 

copies in the reaction mixture in the corresponding amplification microplate well as previously established 

on the Work Sheet. Proceed in this way for the other Q - PCR Standard (10 3 , 10 4 , 10 5 copies). 

9. Carefully seal the amplification microplate using the amplification adhesive sheet. 

10. Transfer the amplification microplate to the real time thermal cycler in the amplification / detection 

area of the amplification products and start the thermal amplification cycle. 

Qualitative analysis of the results 

The values of fluorescence emitted by the specific probe for VZV (FAM fluorescence) and by the 

specific probe for the beta globin (VIC fluorescence) in the amplification reactions must be analysed by the 

instrument software. 

Before analysing, it is necessary to: 

- referring to the instrument documentation, manually set the calculation range for the fluorescence 

back ground level (Baseline) from cycle 6 to cycle 15*; 

*N.B.: in the case of a positive sample with a high titre of VZV, the FAM fluorescence of the specific probe 

for VZV may begin to grow before the 15th cycle. In this case the calculation range for the “baseline” must be 

adjusted from cycle 6 to the cycle in which the FAM fluorescence starts to grow. 

- Referring to the instrument documentation, manually set the Threshold for the FAM fluorescence 

to 0.2; 

- Referring to the instrument documentation, manually set the Threshold for the VIC fluorescence to 

0.1. 

The values of fluorescence emitted by the specific probe for VZV in the Positive control 

amplification reaction and the Threshold value of fluorescence are used to validate amplification and 

detection as shown in the following table: 

VZV Positive control 

Threshold cycle (FAM) 

Assay result 

Amplification / Detection 

Determined POSITIVE CORRECT 

If the result of the Positive control amplification reaction is Undetermined, it means that target 

DNA has not been detected. A problem has occurred during the amplification or detection phase (incorrect 

reaction mixture volumes, probe degradation, positive control degradation, incorrect dispensing of positive 

control, incorrect positive control position setting, incorrect thermal cycle setting) which may cause incorrect 

results. The session is invalid and must be repeated from the amplification phase. 

N.B.: When this product is used for dosing VZV DNA, amplification and detection must be validated in 

relation to the value of fluorescence emitted by the specific probe for VZV in the amplification reactions of the 

four Q - PCR Standards, used instead of the Positive Control, and to the fluorescence Threshold value. 

The values of fluorescence emitted by the specific probe for VZV in the Negative control 

amplification reaction and the Threshold value of fluorescence are used to validate amplification and 

detection as shown in the following table: 

VZV Negative control 

Threshold cycle (FAM) 

Assay result 


Undetermined NEGATIVE CORRECT 

If the result of the Negative control amplification reaction is other than Undetermined, it means that 

target DNA has been detected. Problems have occurred during the amplification phase (contamination) 

which may cause incorrect results and false positives. The session is invalid and must be repeated from the 

amplification phase. 





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RTS035-M 

The values of fluorescence emitted by the probes in the amplification reactions of each sample and 

the Threshold value of fluorescence are used to detect the presence of target DNA and to validate 

amplification and detection by determining the Threshold cycle (Ct), the cycle in which the Threshold value 

for fluorescence is reached. 

This product is able to detect a minimum quantity of 10 DNA copies of the gene that codifies the 

Major DNA binding protein of VZV per amplification reaction, corresponding to the genome Equivalents per 

reaction (see paragraph on Performance Characteristics on page 13). 

When the product is used for detection of VZV, the results for each sample are used as shown in the 

following table: 

Threshold cycle of the sample 

VZV (FAM) beta globin (VIC) 

Undetermined 

Determined 

Ct > 35 or 

Undetermined 

Sample 

suitability 

Assay result 

VZV DNA 

not suitable invalid - 

Ct ≤ 35 suitable valid, negative NOT DETECTED 

Ct > 35 or 

Undetermined 

suitable* valid, positive PRESENT 

Ct ≤ 35 suitable valid, positive PRESENT 

If the result of the amplification reaction of a sample is Undetermined for the VZV DNA and Ct > 35 

or Undetermined for the DNA of the human beta globin gene, this means that it has not been possible to 

sufficiently detect the DNA of the human beta globin gene. In this case, problems have occurred during the 

amplification phase (inefficient or invalid amplification) or in the extraction phase (loss of DNA, presence of 

inhibitors, degradation of the DNA sample or insufficient number of cells in the starting sample), which may 

cause incorrect results and false negatives. The sample is not suitable, the assay is invalid and must be 

repeated beginning with extraction of a new sample. 

If the result of the amplification reaction of a sample is Undetermined for the VZV DNA and Ct ≤ 35 

for the DNA of the human beta globin gene, this means that VZV DNA has not been detected in the DNA 

extracted from the sample but it is not possible to exclude the presence of VZV DNA at a lower titre than the 

detection limit of the product (see paragraph on Performance Characteristics on page 13). In this case the 

result would be a false negative. 

The results obtained with this assay must be interpreted in consideration of all the clinical data and 

the other laboratory tests done on the patient. 

*N.B.: When VZV DNA has been detected in the amplification reaction of a sample, amplification of the 

human beta globin gene may result in Ct > 35 or Undetermined. In fact the low-efficiency amplification 

reaction of the human beta globin gene may be cancelled out by competition with the high-efficiency 

amplification reaction of VZV DNA. In this case the sample is still suitable and the positive result of the assay 

is valid. 

Quantitative analysis of the results 

After carrying out the procedure for qualitative analysis of the results it is possible to perform the 

quantitative analysis of the results of the positive samples. 

The values of fluorescence emitted by the specific probe for VZV in the amplification reactions of the 

four Q - PCR Standard are used to calculate the Standard Curve of the amplification session and to 

validate the amplification and detection as shown in the following table: 

VZV Standard Curve 

(FAM) 

Acceptance range 


Correlation coefficient (R2) 0.990 ≤ R2 ≤ 1.000 CORRECT 

If the Correlation coefficient value (R2) is not within the limits, it means that a problem has 

occurred during the amplification or detection phase (incorrect reaction mixture volumes, probe degradation, 

standard degradation, incorrect dispensing of the standards, incorrect standard position setting, incorrect 

thermal cycle setting) which may cause incorrect results. The session is invalid and must be repeated from 

the amplification phase. 

The values of fluorescence emitted by the specific probe for VZV in the amplification reactions of 

each sample and the Standard Curve of the amplification session are used to calculate the Quantity of 

target DNA present in the amplification reactions of the samples. 

This product is able to dose between 1,000,000 and 10 copies of the DNA of the gene that codifies 

the VZV Major DNA binding protein per amplification reaction, corresponding to the genome Equivalents 

(see paragraph on Performance Characteristics on page 13) as shown in the following table: 

Result of the VZV sample 

(FAM) 

genome Equivalents of VZV per reaction 

Quantity > 1 x 10 6 GREATER THAN 1,000,000 

1 x 10 1 ≤ Quantity ≤ 1 x 10 6 = Quantity 

Quantity < 1 x 10 1 FEWER THAN 10 

When this product is used for measuring of the VZV DNA, the results (Quantity) of the amplification 

reactions of the samples are used to calculate the number of genome Equivalents (gEq) of VZV available in 

the starting sample (Nc) according to this formula: 

Ve x Quantity 

Nc (gEq / ml) = —————————— 

Vc x Va x Ee 

Vc is the volume of the sample used in the extraction; for example with «EXTRAgen ® » the Vc 

parameter is 0.3 ml. 

Ee is the efficiency of the extraction; for example with «EXTRAgen ® » the Ee parameter is 0.8 

(minimum efficiency of 80%). 

Ve is the total volume of the extraction product; for example with «EXTRAgen ® » the Ve parameter is 

15 µl. 

Va is the volume of the extraction product used in the amplification reaction: with this product the Va 

parameter is 5 µl. 

Quantity is the result of the amplification reaction of the sample in gEq. 

When Nanogen Advanced Diagnostics S.r.L. extraction kits are used, the formula becomes: 

Extraction kit 

«EXTRAgen ® » 

Simplified formula 

Nc (gEq / ml) = 12.5 x Quantity / ml 





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Calculation of the dosing range limits 

When a particular extraction assay method is used, the dosing range limits may be calculated from 

the dosing range of the amplification reaction according to the following formula: 

Ve x 10 gEq 

Lower limit (gEq / ml) = —————————— 

Vc x Va x Ee 

Ve x 1,000,000 gEq 

Upper limit (gEq / ml) = —————————— 

Vc x Va x Ee 

When Nanogen Advanced Diagnostics S.r.L. extraction kits are used, the formula becomes: 

Extraction kit 

«EXTRAgen ® » 

Dosing range limits 

from 125 to 12,500,000 gEq / ml 

PROCEDURE LIMITATIONS 

Use only DNA extracted from the following human samples with this product: swabs of 

mucocutaneous lesions, cephalo-rachidian liquid, plasma collected in EDTA. 

Do not use DNA extracted from heparinized samples with this product: heparin inhibits the 

amplification reaction of nucleic acids and causes invalid results. 

Do not use DNA extract that is contaminated with haemoglobin with this product: haemoglobin 

inhibits the amplification reaction of nucleic acids and may cause invalid results. 

There are no data available concerning inhibition caused by antiviral drugs, antibiotics, 

chemotherapeutic drugs or immunosuppressants. 

The results obtained with this product are subject to the correct collection, transport, storage and 

preparation of samples. To avoid incorrect results it is therefore necessary to take particular care during 

these phases and to carefully follow the instructions provided with the products for nucleic acid extraction. 

Owing to its high analytical sensitivity, the real-time amplification assay of nucleic acids used in this 

product is subject to contamination from VZV-positive clinical samples, positive controls and the amplification 

reaction products themselves. Contamination leads to false positive results. The product has been designed 

in such a way as to reduce contamination; nevertheless, this phenomenon can only be prevented by 

following good laboratory practices and by complying scrupulously with the instructions provided in this 

manual. 

This product must be handled by personnel trained in the processing of potentially infective biological 

samples and chemical preparations classified as dangerous to prevent accidents with potentially serious 

consequences for the user and other persons. 

This product requires the use of work clothes and premises that are suitable for the processing of 

potentially infective biological samples and chemical preparations classified as dangerous to prevent 

accidents with potentially serious consequences for the user and other persons. 

This product must be handled by personnel trained in molecular biology techniques, such as 

extraction, amplification and detection of nucleic acids, to avoid incorrect results. 


the amplification/detection of amplification products to prevent false positive results. 

This product requires the use of special clothing and instruments for extraction/preparation of 

amplification reactions and for amplification/detection of amplification products to avoid false positive results. 

A negative result obtained with this product suggests that the VZV DNA was not detected in DNA 

extracted from the sample, but it may also contain VZV DNA at a lower titre than the detection limit for the 

product (see paragraph on Performance Characteristics on page 13); in this case the result would be a false 

negative. 

As with any diagnostic device, the results obtained with this product must be interpreted in 

consideration of all the clinical data and other laboratory tests done on the patient. 

As with any diagnostic device, there is a residual risk of obtaining invalid results, false positives and 

false negatives with this product. This residual risk cannot be eliminated or reduced any further. In particular 

situations such as emergency diagnoses, this residual risk can contribute to incorrect decisions with 

potentially grave consequences for the patient. 





RTS035-M 



RTS035-M 

Analytical sensitivity: detection limit 

PERFORMANCE CHARACTERISTICS 

The analytical sensitivity of this assay enables identification of approx. 10 target DNA molecules in 

5 µl of DNA extract added to the amplification reaction. 

In terms of detection limit, the analytical sensitivity of the assay was tested using plasmid DNA 

containing the amplification product whose initial concentration was measured by spectrophotometer. The 

plasmid DNA was diluted to a titre of 10 copies / 5 µl in human genomic DNA at a titre of 500 ng / 5 µl. This 

sample was used in 50 repeats for amplification with Nanogen Advanced Diagnostics S.r.L. products (see 

paragraph on accessory products). 

The final results are summed up in the following table. 

Samples No. positive negative 

10 copies plasmid DNA + 500 ng of human genomic DNA 50 50 0 

Analytical sensitivity: linear measuring range 

In terms of linear measuring range, the analytical sensitivity of this assay enables determination of a 

titre of between 1,000,000 and 10 target DNA molecules in 5 µl of DNA extract added to the amplification 

reaction. 

In terms of linear measuring range, the analytical sensitivity of the assay was determined using a 

panel of dilutions (1 log10 between one dilution and the next) of plasmid DNA containing the amplification 

product whose initial concentration was measured by spectrophotometer. The panel points from 10 7 

molecules per reaction to 10 1 molecules per reaction were used in 9 repeats for amplification with Nanogen 

Advanced Diagnostics S.r.L. products. 

The analysis of the data obtained, performed via linear regression, demonstrated that the assay has 

a linear response for all the panel points (linear correlation coefficient greater than 0.99). 

The upper limit of the linear measuring range was fixed at 10 6 molecules / 5 µl, within one logarithm 

of the highest concentration Q - PCR Standard amplification standard (10 5 molecules / 5 µl). 

The lower limit of the linear measuring range was fixed at 10 molecules / 5 µl, within one logarithm of 

the lowest concentration Q - PCR Standard amplification standard (10 2 molecules / 5 µl). 


Linear measuring range 

DNA copies / reaction 

gEq / ml 

Upper limit 1.000.000 12.500.000 

Lower limit 10 125 

Analytical sensitivity: Precision 

The assay precision study, that is the variability of the results of different repeats of a sample with 

the same concentration analysed during the same session, made it possible to determine a Coefficient of 

variation (CV %) of 16.8% within the linear measuring range of 10 6 molecules / 5 µl to 10 molecules / 5 µl. 

Analytical sensitivity: Accuracy 

The assay accuracy study, that is the difference between the mean results obtained in a single 

session on different repeats of a sample with the same concentration and the theoretical value of the 

concentration of the samples, made it possible to determine a mean percentage inaccuracy of 10.5% within 

the linear measuring range of 10 6 molecules / 5 µl to 10 molecules / 5 µl. 

Analytical sensitivity: reproducibility with proficiency test panel 

In terms of reproducibility of the results compared with the results obtained using other assays and in 

various laboratories, the analytical sensitivity of the assay was checked via a proficiency test plasma panel. 

The tests were carried out using as the calibrated reference material two panels of VZV dilutions in 

plasma within the concentration limit (QCMD 2003 VZV proficiency panel and QCMD 2004 VZV proficiency 

panel, Qnostics Ltd, United Kingdom). Each panel sample was used in 2 repeats to carry out the entire 

procedure of analysis, extraction, and amplification with Nanogen Advanced Diagnostics S.r.L. products. 

The results are shown in the following table. 

Tests with calibrated reference material QCMD 2003 VZV proficiency panel 

Sample 

Expected result 

Mean results 

Repeat Positive 

(gEq / ml) 

gEq / ml 

VZV1-01 Positive, 3-7 x 10 2 VZV Ellen 2 2/2 4,32 x 10 2 

VZV1-02 Positive, 1-2.3 x 10 2 VZV Ellen 2 2/2 1.65 x 10 2 

VZV1-03 Negative, 3,8 x 10 7 HSV1 2 0/2 Not found 

VZV1-04 Positive, 1-2 x 10 7 VZV SBL 9/84 2 2/2 0.70 x 10 7 

VZV1-05 Positive, 3.3-6.7 x 10 2 VZV SBL 9/84 2 2/2 2.61 x 10 2 

VZV1-06 Positive, 3-7 x 10 3 VZV Ellen 2 2/2 2.98 x 10 3 


VZV1-08 Negative 2 0/2 Not found 





Tests with calibrated reference material QCMD 2004VZV proficiency panel 

Sample 

Expected result 

Mean results 

Repeat Positive 

(gEq / ml) 

gEq / ml 

VZV04-01 Positive, 3.3 x 10 4 VZV Ellen 2 2/2 1,7x10 4 

VZV04-02 Positive, 7.0 x 10 6 VZV SBL 9/84 2 2/2 2.9x10 6 




VZV04-06 Negative, 2.1 x 10 8 HSV1 2 0/2 Not found 

VZV04-07 Positive, 1.1 x 10 3 VZV Ellen 2 2/2 0.6x10 3 

VZV04-08 Positive, 3.3 x 10 6 VZV Ellen 2 2/2 1x10 6 

VZV04-09 Positive, 3.3 x 10 3 VZV Ellen 2 2/2 1.6x10 3 


The product was capable of correctly reproducing the results obtained with other assays and in 

different laboratories. 

In these tests, a gEq of Qnostics Ltd. was equivalent to approx. 0.5 gEq calculated with 

«VZV Q - PCR Standard» by Nanogen Advanced Diagnostics S.r.L. 





RTS035-M 



RTS035-M 

Diagnostic sensitivity: efficiency of detection and quantification on different genotypes / subtypes 

The diagnostic sensitivity of the assay, that is the efficiency of detection and quantification on 

different genotypes / subtypes, was evaluated by comparison of sequences with nucleotide databases. 

The alignment test of the regions chosen for hybridization of the AmpliMIX primer oligonucleotides 

and of the AmpliPROBE fluorescent probe with the sequences available in the database of the gene 

codifying the Major DNA binding protein of VZV showed preservation and absence of significant mutations. 

The diagnostic sensitivity of the assay, that is the efficiency of detection and quantification on 

different genotypes / subtypes, was evaluated by analysing samples that were positive for DNA of two 

strains of VZV. 

The diagnostic sensitivity of the assay was evaluated using, as the reference material, samples 

certified positive for the Ellen strain of VZV DNA, ATCC, and SBL 9/84 strain of VZV, SMI, (QCMD 2003 VZV 

proficiency panel and QCMD 2004 VZV proficiency panel, Qnostics Ltd, United Kingdom). Each sample was 

used to carry out the entire procedure for analysis, extraction and amplification with Nanogen Advanced 

Diagnostics S.r.L. products. The results are shown in the following table: "Analytical sensitivity: 

reproducibility with proficiency test panel”. 

Diagnostic sensitivity: positive samples 

The diagnostic sensitivity of the assay, confirming positive clinical samples, was evaluated by 

analysing some clinical samples that were positive for VZV DNA and was greater than 81.2%. 

The diagnostic sensitivity was evaluated using 11 mucocutaneous swabs and 6 cephalo-rachidian 

liquid samples that were positive for VZV DNA as the reference material (tested using the Nested 

Amplification assay). Each sample was used to carry out the entire procedure for analysis, extraction and 

amplification with Nanogen Advanced Diagnostics S.r.L. products. 



Mucocutaneous swabs positive for VZV DNA 11 10 1 

Cephalo-rachidian liquid positive for VZV DNA 6 3 2 

A mucocutaneous swab sample and two samples of cephalo-rachidian liquid produced a conflicting 

negative result using the Nanogen Advanced Diagnostics S.r.L. products. These samples were tested again 

using the nested amplification assay and gave a negative result. The conflict can be explained by a titre of 

VZV DNA below the detection limit of the two assays, which produces random positive results or by VZV 

DNA degradation in the starting sample. 

A sample of cephalo-rachidian liquid was invalid. 

Analytical specificity: Potential interference markers 

The analytical specificity of the assay, that is the cross-reactivity with other potential interference 

markers, was evaluated by comparison of sequences with nucleotide databases. 

The alignment test of the regions chosen for hybridization of the AmpliMIX primer oligonucleotides 

and of the AmpliPROBE fluorescent probe with the sequences available in databases of organisms other 

than VZV, including the complete HSV1 and HSV2 genome, the human herpetic viruses that are most similar 

to VZV, showed their specificity and the absence of significant homology. 

In terms of absence of cross-reactivity with other potential interference markers, the analytical 

sensitivity of the assay was evaluated by analysing some samples that were HSV1 and HSV2 DNA-positive. 

The analytical specificity of the assay was tested using, as the reference material, high-titre dilutions 

of HSV1 and HSV2 DNA in plasma (PeliCheck HSV-1/2-DNA-02, AcroMetrix Europe B.V., Netherlands). 

Each sample was used to carry out the entire procedure for analysis, extraction and amplification with 

Nanogen Advanced Diagnostics S.r.L. products. 



Positive samples for HSV1 DNA 1 0 1 

Positive samples for HSV2 DNA 1 0 1 

Diagnostic specificity: negative samples 

The diagnostic specificity of the assay, confirming negative clinical samples, was tested by analysing 

a panel of normal donor plasma samples and a number of VZV DNA-negative samples and proved to be 

equal to 97.7%. 

The diagnostic specificity was evaluated using 20 normal donor plasma samples as reference 

material (Panel Normal Human Plasma, Acrometrix Europe B.V., the Netherlands). Each panel sample was 

used to carry out the entire procedure for analysis, extraction and amplification with Nanogen Advanced 

Diagnostics S.r.L. products. 



Normal donor plasma 20 0 20 

The diagnostic specificity was evaluated using 12 mucocutaneous swabs and 13 cephalo-rachidian 

liquid samples that were positive for VZV DNA as the reference material (tested using the Nested 

Amplification assay). Each panel sample was used to carry out the entire procedure for analysis, extraction 

and amplification with Nanogen Advanced Diagnostics S.r.L. products. 



Mucocutaneous swabs negative for VZV DNA 12 1 11 

Cephalo-rachidian liquid negative for VZV DNA 13 0 12 

A mucocutaneous swab sample produced a conflicting positive result using the Nanogen Advanced 

Diagnostics S.r.L. products. The conflict can be explained by a very low titre of VZV DNA (less than 10 gEq / 

reaction) which is below the detection limit (that is it produces random positive results) of the nested 

amplification system used to test the samples. 

A sample of cephalo-rachidian liquid was invalid. 

N.B.: The complete data and results of the tests carried out to evaluate the performance characteristics of 

the product are recorded in Section 7 of the Product Technical File "VZV Q - PCR Alert AmpliMIX" and 

"VZV Q - PCR Alert AmpliPROBE", FTP RTS035. 

REFERENCES 

A. J. Wakefield et al. (1992) J Med Virology 38: 183 - 190 





RTS035-M 



RTS035-M 

TROUBLESHOOTING 

SYMBOLS 

Target DNA not detected in the Positive Control / Q - PCR Standard reaction or 

invalid correlation coefficient of the standard curve. 


Possible causes 

Error in the preparation of the reaction mixture. 

Dispensing error on the microplate. 

Probe degradation. 

Solutions 

Check the volumes of reagent dispensed during 


Take care when dispensing reactions onto the microplate 

and comply with the work sheet. 

Check the volumes of reaction mixture dispensed. 

Check the volumes of standard dispensed. 

Use a new probe aliquot. 

Upper temperature limit. 

Batch code. 


Positive control or standard degradation. 

Instrument setting error. 

Target DNA detected in the negative control reaction 

Use a new aliquot of Positive control or standard. 

Check the position settings for the positive control or 

standard reactions on the instrument. 

Check the thermal cycle settings on the instrument. 





Dispensing error on the microplate. 

Error while setting the instrument 

Microplate badly sealed. 

Contamination of the sterile bidistilled water. 

Contamination of the amplification mix. 

Solutions 

Avoid spilling the contents of the sample test tube. 

Always change tips between one sample and another. 

Take care when dispensing samples, negative controls, 

positive controls and standards onto the microplate and 

comply with the work sheet. 

Check the position settings of the samples, negative 

controls, positive controls and standards on the instrument 

Take care when sealing the microplate. 

Use a new aliquot of sterile water. 

Use a new aliquot of amplification mix. 



Manufacturer. 

Contamination of the extraction/preparation 

area for amplification reactions. 

High levels of background fluorescence in the reactions 

Clean surfaces and instruments with aqueous detergents, 

wash lab coats, replace test tubes and tips in use. 

Baseline setting error. 


Solutions 

Set baseline calculation interval within cycles where the 

background fluorescence has already stabilized (check the 

"component" recordings) and where the signal 

fluorescence has not started to grow yet, e.g. from cycle 9 

to cycle 15. 



licensed products for "Positive Control" or "Q - PCR Standard". 







VZV Q - PCR Alert AmpliPROBE 


RTS035-P 

«DUPLEX REAL-TIME AMPLIFICATION» 



RTS035-P 

Offices: 

Tel. +39-011 976 19 1 

Fax +39-011 936 76 11 

E. mail: techsupport@nanogenad.com 










REFERENCES page 4 


INTENDED USE 

«VZV Q - PCR Alert AmpliPROBE» is part of a quantitative amplification assay of nucleic acids for 

the detection and dosing of the DNA of herpetic Varicella Zoster virus (VZV) in DNA samples extracted 

from swabs of mucocutaneous lesions, cephalo-rachidian liquid and from plasma collected in EDTA. 

The product is intended for use, alongside clinical data and other laboratory tests, in the diagnosis 

and monitoring of VZV infections. 


The product supplies the mixture of AmpliPROBE fluorescent probes for real time amplification in a 

stabilizing solution, pre-dosed in aliquots into four disposable test tubes. Each test tube contains 110 µl 

of solution, sufficient for 24 reactions. 

The VZV probe, labelled with FAM fluorophore and blocked by the MGB-NFQ group, is specific for a 

region of the gene that codifies the major DNA binding protein of VZV (ORF 29). 

The probe for the internal suitability test of the sample, labelled with VIC fluorophore and blocked by 

the MGB-NFQ group, is specific for the promoter region and 5’ UTR of human beta globin gene. 

The procedure involves a real-time amplification reaction on a microplate with programmable heater 

with optical fluorescence detection system (thermal cycler for real time). 

System standardization was carried out on Applied Biosystems ABI PRISM TM 7000 series 

instruments. 

The kit provides 96 determinations, including standards and controls. 

-20°C 



VZV AmpliPROBE 

Mixture of fluorescent probes 

labelled with FAM / MGB-NFQ 

and VIC / MGB-NFQ 

4 x 110 µl 

Fluorescent oligonucleotides, 


Glycerol, Triton X-100 




- Vortex mixer. 

- Bench microcentrifuge (12,000 – 14,000 RPM). 


50-200 µl, 200-1000 µl). 


- Programmable heater with optical fluorescence detection system. 


The reagents for DNA extraction from the samples to be analysed, the positive extraction control, the 

reagents optimized for amplification, the primer reagents (oligonucleotides), the positive amplification control 

or the known-quantity DNA standard are not included in this product. To perform these analytical steps the 

following accessory products, manufactured by Nanogen Advanced Diagnostics S.r.L., are recommended: 

«EXTRAgen ® » (code EXTG01), kit for extraction of nucleic acids from non-cellular samples; the kit 

enables 50 extractions. 

«CPE-DNA ® - Internal Control» (code CTREXTG), positive plasmid DNA extraction control for noncellular 

sample DNA extractions; the product enables 50 extractions. 

«Q - PCR Alert AmpliMASTER» (code RTS000), combination of optimized reagents, microplates 

and adhesive sheets for real time amplification; the product enables 96 reactions. 

« VZV Q - PCR Alert AmpliMIX» (code RTS035-M), primer oligonucleotides for real time 

amplification; the product enables 96 reactions. 

If a qualitative result of the analysis is required (detection of VZV DNA): 

«VZV - Positive Control» (code CTR035), positive amplification control of plasmid DNA; the product 

enables 25 sessions. 

If a quantitative result of the analysis is required (dosing of VZV DNA): 

«VZV Q - PCR Standard» (code STD035), known-quantity plasmid DNA to obtain the standard 

curve; the product enables 16 sessions. 








Handle and dispose of all reagents and all assay materials as if they were potentially infective. Avoid 

direct contact with the reagents. Avoid splashing or spraying. Waste must be treated and disposed of in 

compliance with the appropriate safety standards. Disposable combustible materials must be incinerated. 

Liquid waste containing acids or bases must be neutralised before disposal. 

- 

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RTS035-P 



RTS035-P 




































Warnings and precautions specific to reagents 

The test tubes containing AmpliPROBE are disposable and therefore must be used once only in the 


The AmpliPROBE carries the following safety warnings (S): 


PROCEDURE 

The product «VZV Q - PCR Alert AmpliPROBE» must be used with the products 

«Q - PCR Alert AmpliMASTER» and «VZV Q - PCR Alert AmpliMIX» to obtain the reaction mixture. 

AmpliPROBE is ready for use, hence must be added directly to the reaction mixture. 

The complete procedure involves preparation and execution of a real time amplification reaction on a 

microplate with programmable heater with optical fluorescence detection system (thermal cycler for real time) 

and is described in detail in the instruction manual enclosed with the «VZV Q - PCR Alert AmpliMIX» 

product. 

The performance characteristics and procedure limitations of the complete assay for detection and 

dosing of VZV DNA are described in detail in the instruction manual enclosed with the 

«Q - PCR Alert AmpliMIX» product. 


REFERENCES 

A. J. Wakefield et al. (1992) J Med Virology 38: 183 - 190 

Upper temperature limit. 

Batch code. 



SYMBOLS 





Manufacturer. 



licensed products for "Positive Control" or "Q - PCR Standard". 


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