The peripheral-type benzodiazepine receptor in rat kidney has been identified by photoaffinity labeling. PK 14105, a derivative of the selective peripheral-type ligand PK 11195, was used to covalently label peripheral-type benzodiazepine receptors. In the absence of UV light PK 14105 demonstrated reversible, high affinity (KD = 4.8 nM) binding to rat kidney mitochondrial membranes. Inhibition of the reversible binding of [3H]PK 14105 by various benzodiazepine and other ligands demonstrated that this ligand bound with all the characteristics expected of a ligand interacting specifically with peripheral-type benzodiazepine receptors. A similar order of relative potencies was obtained for inhibition of photolabeling, indicating that reversible binding and photolabeling occurred at the same class of binding sites. Examination of photolabeled binding sites from kidney, heart, brain and adrenal membranes using sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the probe is photoincorporated into a single peptide of Mr = 18,500. The results indicate that [3H]PK 14105 identifies the ligand binding domain of the peripheral-type benzodiazepine receptor, which is a peptide with Mr = 18,500, that is of similar size in kidney, heart, brain and adrenals.