Abstract
Epstein-Barr Virus (EBV) is a B-lymphotropic herpesvirus which the majority of adult human population is latently-infected with. Various immunological and molecular in vitro studies have been facilitated by the use of EBV’s ability to infect and transform B cells to immortalized polyclonal B cell lines. Many of these studies use freshly isolated cord-blood mononuclear cells (CBMC). Some experiments may, however, require EBV infection of samples that have been prospectively collected and cryo-preserved. Here we share a protocol that we used to successfully infect B cells from cryo-preserved CBMCs and peripheral-blood mononuclear cells (PBMC) (Sohlberg et al., 2013; Saghafian-Hedengren et al., 2013).
Materials and Reagents
Equipment
Procedure
Note about sample handling: Gentle and fast processing of samples during isolation of mononuclear cells contributes to higher B cell viability, which is central for successful in vitro infection.
Representative data
Refer to Figure 1 in Sohlberg et al. (2013) for representative data on B cell characteristics following in vitro EBV infection of B cells.
Notes
Recipes
Acknowledgments
This protocol has been adapted from the previously published paper Sohlberg et al., (2013). At the time of development and implementation of this protocol, the following sources of funding were used: Swedish Research Council Grants 57X-15160-07-03 and 57X-15160-10-4, the Swedish Association for Allergology, the Ragnar Söderberg Foundation, the Ellen, Walter and Lennart Hesselman Foundation, the Konsul Th. C. Bergh Foundation, the Golden Jubilee Memorial Foundation, the Petrus and Augusta Hedlund Foundation, the Crown Princess Lovisa’s/Axel Tielman’s Foundation, the Swedish Cancer Society, and the Karolinska Institute. E.R. and N.N. are recipients of cancer research fellowships from the Cancer Research Institute (New York)/Concern Foundation (Los Angeles). We thank Ehsan Rasul and Eva Klein (Department of Microbiology, Tumor and Cell Biology, KI, Stockholm, Sweden) for their contribution in optimizing the protocol.
References
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