B9 Cell Line mouse

Mouse B cell hybridoma, IL-6 assay

B9 is a mouse B cell hybridoma line that requires Interleukin 6 (IL-6) for survival and proliferation. IL-6 is a pluripotent cytokine with multiple effects on many different cell types. It is produced by a variety of cells in response to various stimuli. The B9 hybridoma allows for a simple sensitive assay for human, rat, rabbit, pig and mouse IL-6 based on the IL-6 dependent proliferation of this murine B cell line. Details of the B9 hybridoma growth factor assay can be found in Current protocols in Immunology 1996 Cytokines and Their Cellular receptors 6.6.1-6.6.5 contributed by Nordan RP, Richards CD and Gauldie J. John Wiley & Sons, Inc.

IDMEM or RPMI 1640 + 2mM Glutamine + 50μM 2-Mercaptoethanol (2ME) + 50pg/ml recombinant human IL-6 (Sigma SRP3096, diluted and stored in PBS containing 1% BSA) + 5% FBS. IL-6 needs to be added fresh at each media change.

For B9 cell maintenance cells should be seeded at 0.5 x 10⁵ cells/ml in B9 culture medium. The culture will be in log phase growth between days 3-5. When the culture reaches 1 x 10⁶ cells/ml (4-5 days) the culture must be sub cultured as above again seeding at 0.5 x 10⁵ cells/ml.; 5% CO₂; 37°C. It maybe necessary to determine the activity of the IL-6 or IL-6 containing supernatant first. The lowest concentration at which maximal growth can be obtained should be used. A fresh vial of cells is required when the background response from control cultures begins to rise. It is important that the culture conditions are adhered to avoid the B9 cells becoming IL-6 independent.

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