Question

You have a viral stock of an unknown concentration and you want to determine the titer of the virus by plaque assay. Your lab mate has VERO cells growing in the lab and you decide to use them for a plaque assay. After infection with a range of dilutions, none of the plates showed plaques. Give 2 possible reasons why you did not get any plaques.

You repeat the plaque assay using CHO as the target cells this time. You infect 3 wells with 0.1 ml of a 10-6 dilution of your stock. The plates show 105, 95 and 100 plaques. What is the titer of your undiluted viral stock?

Answer 1

During plaque assay of virus on VERO cells, no plaques were formed. Two possible reasons for non-occurence of the plaques maybe:

1. The virus may not be infecting the vero cells i.e., Vero cells are kidney cells and hence the virus doesnot infect the kidney cells and hence there is no plaques that are observed.

2. The viral titer in the dilutions maybe low or else the virus stock has lost its infectivity capacity due to long term storage.

Detection of viral titer in plaque assay is calculated as plaque forming units(PFU)/ml and the formula used is:

PFU/ml = number of plaques counted/(dilution*volume of virus culture added)

number of plaques counted = average of all the plaques in the 3 wells = 105+95+100/3 = 100

dilution = 10-6

volume of virus culture added = 0.1ml

PFU/ml = 100/10-6*0.1 = 100*107PFU/ml

Hence this is the viral titer of the virus when plaque assay is conducted on CHO cells.

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