Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrin 𝛽1

Figure 1

Formaldehyde cross-linking. (a) Workflow of formaldehyde cross-linking. Cells are treated with formaldehyde, lysed and protein complexes (oval shapes) are precipitated by antibodies (𝑦-shaped). Cross-links are indicated by black triangles. Only antibodies, whose epitopes are not destroyed during formaldehyde modification, can precipitate the complex. (b) Reaction scheme of formaldehyde modification, cross-linking and cross-linking reversal. (c) and (d) Formaldehyde derived cross-links are preserved, if samples are only incubated at 65^∘C, whereas most of the cross-links are reversed at 99^∘C. (c) Schematic model. Proteins are depicted as oval shapes, formaldehyde cross-links as black triangles. (d) Anti-integrin 𝛽1 immunoblot analysis of cross-linked Jurkat cells (JB1A, anti-mouse HRP).