The cross-linker dithiobis(succinimidyl propionate) (DSP; Thermo Fisher Scientific no. 22586) was freshly prepared at 20 mM in dimethyl sulfoxide and diluted to a working concentration of 0.5 mM in PBS. Cells were washed twice with PBS and incubated in 0.5 mM DSP for 30 min at room temperature. After removal of DSP, cells were incubated with quenching solution [20 mM tris-Cl (pH 7.4)] for 15 min at room temperature, lysed, and incubated with anti-TRAPα antibodies for 6 hours at 4°C. Immunoprecipitates were washed, subjected to SDS-PAGE, transferred to nitrocellulose, and immunoblotted with anti-TRAPα and anti-proinsulin antibodies.

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