FTD Respiratory pathogens 21 plus - Mikrogen

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FTD Respiratory pathogens 21 plus version 5/ March 2012 english FTD Respiratory pathogens 21 plus 11.2 Main PCR setup Procedure with using the AgPath One-Step RT- PCR Kit If you want to use a different enzyme to the AgPath-ID One-Step RT-PCR Kit please firstly check with FTD! Preparation of PCR with AgPath-ID One-Step RT-PCR Kit: 1. Thaw reagents for the reaction: FluRhino PPmix, Cor PPmix, Para.EAV PPmix, Bo.Mp.Pf1 PPmix, RsEPA PPmix and RespBac PPmix, the positive control (Resp PC and RespBac PC) and the 2xRT PCR buffer of AgPath- ID One-Step RT-PCR Kit. The PC`s and the extracted NC have to be included in each run. Make sure to keep the enzyme of the AgPath-ID One-Step RT-PCR Kit cool. It should be kept in a freezer or on a cooling block at all times! 2. Reagents in each column are sufficient for: FTD 2+-24/4 4 patients, including controls FTD 2+-48/6 6 patients, including controls FTD 2+-96/12 12 patients, including controls 3. Pipette the required amount (see table 2 below) of the “2xRT PCR buffer” (AgPath-ID One-Step RT-PCR Kit) to the FluRhino PPmix, Cor PPmix, Para.EAV PPmix, Bo.Mp.Pf1 PPmix, RsEPA PPmix and RespBac PPmix. Take care to change the tips after each pipetting step. 4. Pipette the required amount (see table 2 below) of the “25x RT-PCR enzyme mix” (AgPath-ID One-Step RT-PCR Kit) to FluRhino PPmix, Cor PPmix, Para.EAV PPmix, Bo.Mp.Pf1 PPmix, RsEPA PPmix and RespBac PPmix with the “2xRT PCR buffer”. Take care to change the tips after each pipetting step. Vortex the complete master mixes briefly and spin down in a centrifuge. - 12 -
FTD Respiratory pathogens 21 plus version 5/ March 2012 english FTD Respiratory pathogens 21 plus Table 2: Shown are the amounts of reagents that are needed for one well and for multiples accordant to the kit size. FTD 2+-24/4: The PPmix contained in one tube is sufficient for 7 reactions and includes 4 patients, 1 NC, 2 PC, 10 % pipetting inaccuracy. FTD 2+-48/6: The PPmix contained in one tube is sufficient for 9 reactions and includes 6 patients, 1 NC, 2 PC, 10 % pipetting inaccuracy. FTD 2+-96/12: The PPmix contained in one tube is sufficient for 15.5 reactions and includes 12 patients, 1 NC, 2 PC, 10 % pipetting inaccuracy. FTD 2+ -24/4 FTD 2+ -48/6 FTD 2+ -96/12 Number of samples 1 7 PP mix 1.5 µl 10.5 µl Preparation of the PCR assay - 13 - Enzyme 1 µl 7 µl Buffer 12.5 µl 87.5 µl Total 15 µl 105 µl Number of samples 1 9 PP mix 1.5 µl 13.5 µl Preparation of the PCR assay Enzyme 1 µl 9 µl Buffer 12.5 µl 112.5 µl Total 15 µl 135 µl Number of samples 1 15.5 PP mix 1.5 µl 23.3 µl Preparation of the PCR assay Enzyme 1 µl 15.5 µl Buffer 12.5 µl 194 µl Total 15 µl 232.8 µl
Page 1 and 2: FTD Respiratory pathogens 21 plus v
Page 3 and 4: 1. Identification of the manufactur
Page 7 and 8: 5. Contents FTD Respiratory pathoge
Page 9 and 10: 8. Principle of the method FTD Resp
Page 11: 11. Procedure FTD Respiratory patho
Page 15 and 16: Preparation of a 96 well plate for
Page 17 and 18: 12. Programming of the thermo cycle
Page 25 and 26: 17. Troubleshooting FTD Respiratory
Page 27 and 28: There are only two exceptional case
Page 31 and 32: Pathogen FluA FTD resp+ H1N1 FTD Fl

FTD Respiratory pathogens 21 plus - Mikrogen

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