Methods to Study the Role of Cdc42, Rac1, and Rac2 in B-Cell Cytoskeletal Responses
B-cell migration and adhesion are critical to form a germinal center response, the site for B-cell production of high-affinity antibodies. Here, we describe two assays that can be used to examine B-cell cytoskeletal responses needed during the germinal center response: B-cell spreading and homotypic adhesion. Spreading of B cells is dependent on Cdc42, while Rac1 and Rac2 are necessary for homotypic adhesion. These in vitro assays can be used to examine functional responses of B cells mediated by the cell cytoskeleton, for example when comparing B cells from different gene knockout animals.
Key wordsB lymphocytes In vitro responses Polyclonal activation Adhesion Spreading
Formation of aggregates in B cells cultured with anti-CD40 + IL-4. B cells were stimulated for 2 days. Bright-field images were taken every minute. Note that big aggregates are formed by fusion of smaller ones. Magnification 10× (MOV 61702 kb)
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