For the first time I got larger plaques for PR8 while later i got smaller plaques. The TPCK trypsin I used were prepared at different times.
Dear Prem, As plaque is a localized area of CPE and TPCK trypsin promote the internalization of the virus and the growth of CPE. so, different permissible concentrations of TPCK trypsin based on the subtype of influenza virus and cell line may affect the plaque size formation.
regards
Was the same batch of TPCK treated trypsine used both times? The units active enzyme per mg might differ between batches, hence resulting in a different concentration and subsequently having an effect on the amount of cleaved HA0 and as a result the amount of infectious virus. Each batch TPCK treated trypsin should be titrated for similar results.
Of course it does. In addition, TPCK trypsin is sensitive to repeated freeze-thawing (so it is better to aliquot it and to use only fresh TPCK). In contrast, many cell lines are highly sensitive to higher concentrations of TPCK trypsin and for some cell lines it is necessary to use lower concentrations than classically used with MDCK cells for instance.
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