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- DocumentStandard Nested PCR Protocoluploaded byKris Cahyo Mulyatno
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- DocumentStandard Nested PCR Protocoluploaded byKris Cahyo Mulyatno
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- DocumentVirus titration by using the plaque assay 1. Prepare confluent monolayers of cells (vero cell) in 24 well plates. 2. Prepare serial 10-fold dilutions (101 to 107 ) of virus in chilled maintenance medium (MEM, with 1% serum). 3. Remove culture medium and add 0.2ml of virus inoculum, starting from the highest dilution. Ensure that a film of medium completely covers the cell sheet. 4. Incubate the plate at 37 C for 1 hour with intermittent rocking of the plate. 5. Remove the inoculum, preferably with a pipette and then add 1.5 ml of agarose overlay medium (growth medium with 0.3% agarose and 2.5% FCS). 6. Ensure that the overlay medium has spread evenly over the monolayer, leave at room temperature for 10 mins and then incubate at 37 C . 7. Examine the monolayers daily, starting from second day of incubation. 8. Once the plaques have developed, usuallyuploaded byKris Cahyo Mulyatno
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