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. 1999 Oct 18;147(2):401–416. doi: 10.1083/jcb.147.2.401

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© 1999 The Rockefeller University Press

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).

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Three-dimensional collagenous matrix fails to activate FAK in Saos-2 cells. Cells from two separate α2 or α2/α1 single cell clones (45, 47 and 2, 12) were pooled together and either lysed immediately (0 h sample), seeded in collagen gel, or allowed to adhere to fibronectin for 1 h. FAK was immunoprecipitated and immunoplotted with an antiphosphotyrosine antibody (4G10). Alternatively, cell lysates were fractionated on SDS-PAGE and immunoblotted with phosphospecific FAK antibody. The blots were reprobed with anti–FAK antibody to show the levels of FAK protein. Note that in the lower panel the anti-phospho FAK antibody recognizes two bands, but it is the upper one that comigrates with the band recognized with the anti–FAK antibody. The origin of the lower band is unknown.