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. 1997 Dec 15;139(6):1455–1464. doi: 10.1083/jcb.139.6.1455

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sTva dissociates from API. (A) sTva or sTva-API was incubated with liposomes and subjected to centrifugation as in Fig. 1. Gradient fractions were chloroform-methanol precipitated, separated by SDS-12.5% PAGE, transferred to nitrocellulose, and Western blotted with anti-Tva serum. (B) Unlabeled API was immunoprecipitated with the anti-DAF antibody and protein A agarose. Biotinylated sTva was added to the immunoprecipitates on ice for 20 min. Liposomes or PBS were added and samples were incubated for 5 min at 37°C. Beads were centrifuged and the supernatants collected and chloroform-methanol precipitated. Beads and supernatant proteins were reduced, boiled, separated by SDS-12.5% PAGE, transferred to nitrocellulose, and blotted with streptavidin-HRP.

sTva dissociates from API. (A) sTva or sTva-API was incubated with liposomes and subjected to centrifugation as in Fig. 1. Gradient fractions were chloroform-methanol precipitated, separated by SDS-12.5% PAGE, transferred to nitrocellulose, and Western blotted with anti-Tva serum. (B) Unlabeled API was immunoprecipitated with the anti-DAF antibody and protein A agarose. Biotinylated sTva was added to the immunoprecipitates on ice for 20 min. Liposomes or PBS were added and samples were incubated for 5 min at 37°C. Beads were centrifuged and the supernatants collected and chloroform-methanol precipitated. Beads and supernatant proteins were reduced, boiled, separated by SDS-12.5% PAGE, transferred to nitrocellulose, and blotted with streptavidin-HRP.