Hallo everyone,
I have a problem with plasma toxicity in cell cultures during virus titrations.
More specifically, we infect CD-1 mice with a virus, which we believe is capable of causing a transient viremia. The virus grows perfectly in cell cultures, so high doses can be used to infect mices.
Then we collect blood plasma from the infected mice at various time intervals, to measure the amounts of the virus in the plasma (viremia). The collected plasma is EDTA-stabilized.
Then we do virus titration experiments in BHK-21 cells.
However, the plasma at small dilutions (1:10, 1:100) exerts a strong cytopathic effect on BHK-21 cells. This cytopathic effect is not connected to the infection because the same cell killing can be seen if plasma from uninfected mice is used as a control. There is no CPE is the plasma is diluted to greater dilutions (1:1000...).
I did not have observed such CPE with sera from different sources. This is possibly an effcet of complement, however I did not checked.
I am afraid the CPE (in wells with small plasma dilutions) interferes with the correct measuring of virus titers if the titers are small.
1) What can I do to supress the nonspecific CPE (plasma toxicity) in small dilutions preserving the virus infection. Obviously I cannot heat the plasma for complement inctivation.
2) Is blood sera more suitable material to measure viremia with less pronounced toxicity to cell cultures compared to plasma?
3) Have any person experience a similar problem?
How can I measure a small-titers viremia if the nonspecific plasma toxicity is present in my experiments? I don want to swith to molecular or indirect methods such as PCR, ELISA etc. I want to measure the virus by infectivity.
Thanks everyone for true expertize.
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