In vitro import experiments with isolated organelles are a powerful tool for investigation of the biogenesis of proteins. A key issue in such experiments is an assay to distinguish between correctly and incorrectly imported proteins. Here we describe an assay to monitor in vitro the proper membrane integration of single-span proteins. In this assay non-imported proteins are distinguished from correctly imported protein species by labelling of unprotected cysteine residues and a resulting migration shift in SDS-PAGE.
