I am trying to adopt a rotavirus from clinical samples to MA-104 cell line, the virus starts producing the CPE from the second passage and virus presence was confirmed by ELISA and RT-PCR from both first and second passage, but from second passage onwards the virus titre is reduced and in some cases it disappeared (fails to produce CPE and couldn't be detected both by ELISA and RT-PCR). I have tried multiples times from back passages with some modification in Trypsin concentration but it hasn't worked out. Can any one explain to me what could the reason and the best method to adopt rotavirus to cell culture from clinical samples?
Typically, clinical samples are adapted to growth in primary AGMKs and secondary AGMKs, before transitioning to MA104 cells. You might check the following reference:
Arnold M, Patton JT, McDonald SM. Culturing, storage, and quantification of
rotaviruses. Curr Protoc Microbiol. 2009 Nov;Chapter 15:Unit 15C.3. PubMed PMID:
19885940.
National Institute of Virology, Pune, has a lot of experience in isolating rotaviruses from clinical samples. The institute may even have a good number of isolates that are already adapted to growth in cell culture.
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