Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrin 1
Figure 4
Precipitation of cross-linked integrin complexes. (a, b) Jurkat cells were treated with or without 0.4% formaldehyde (PFA) before lysis. Immunoprecipitation was performed using 0.5 mg of lysate and 1 g of the indicated antibody. Immunoprecipitations (IP) and supernatants (SN) were analyzed for presence of integrin complexes using JB1A for immunoblotting (anti-mouse Alexa Fluor 680). (a) JB1B precipitated the integrin complex (indicated by arrow) from formaldehyde treated cells. (b) Test of the remaining seven antibodies for precipitation of integrin from formaldehyde treated cells using the same parameters as in (a). (c) Jurkat cells were treated with different concentrations of formaldehyde (PFA) and lysed. 100 g lysate were used for pull-downs with JB1B (1 g) and immunoprecipitations (IP) and supernatants (SN) were analyzed by anti-integrin immunoblotting (JB1A, anti-mouse HRP).
