2015
Human norovirus culture in B cells
Publication
Publication
Nature Protocols , Volume 10 - Issue 12 p. 1939- 1947
Human noroviruses (HuNoVs) are a leading cause of foodborne disease and severe childhood diarrhea, and they cause a majority of the gastroenteritis outbreaks worldwide. However, the development of effective and long-lasting HuNoV vaccines and therapeutics has been greatly hindered by their uncultivability. We recently demonstrated that a HuNoV replicates in human B cells, and that commensal bacteria serve as a cofactor for this infection. In this protocol, we provide detailed methods for culturing the GII.4-Sydney HuNoV strain directly in human B cells, and in a coculture system in which the virus must cross a confluent epithelial barrier to access underlying B cells. We also describe methods for bacterial stimulation of HuNoV B cell infection and for measuring viral attachment to the surface of B cells. Finally, we highlight variables that contribute to the efficiency of viral replication in this system. Infection assays require 3 d and attachment assays require 3 h. Analysis of infection or attachment samples, including RNA extraction and RT-qPCR, requires ∼6 h.
Additional Metadata | |
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doi.org/10.1038/nprot.2015.121, hdl.handle.net/1765/91500 | |
Nature Protocols | |
Organisation | Department of Virology |
Jones, M. K., Grau, K. R., Costantini, V., Kolawole, A. O., de Graaf, M., Freiden, P., … Karst, S. M. (2015). Human norovirus culture in B cells. Nature Protocols, 10(12), 1939–1947. doi:10.1038/nprot.2015.121 |