Two derivatives of the peptide hormone cholecystokinin (CCK-8s) containing a photocleavable o-nitrobenzylester group have been synthesized. One analog, [3H]4-(biotin-epsilon-Ahx-oxymethyl)-3-nitrobenzoyl-Gly-Orn (propionyl)-epsilon-Ahx-CCK-8s ([3H]BANA-CCK-8s), was biotinylated, while the other, 4-alanyloxymethyl-3-nitrobenzoyl-epsilon-Ahx-CCK-8s (ANA-CCK-8s) had a free amino group for coupling to amino-reactive affinity matrices. The analogs retained high affinity to both CCK receptors and anti-CCK antibodies. [3H]BANA-CCK-8s was bound to a streptavidin-agarose affinity matrix and was subsequently released by irradiation with uv light of wavelengths > 320 nm. The ability to recover a biotinylated peptide ligand from a streptavidin column under very mild conditions should be useful in the purification of specifically binding proteins. As a demonstration of this approach, [3H]BANA-CCK-8s was incubated with anti-CCK-antiserum and was subsequently passed over a streptavidin-agarose affinity matrix. After washing, the bound antibodies were eluted by photocleavage of the affinity ligand. Forty-six percent of the bound antibodies were recovered after 40 min of irradiation. The eluted antibodies showed essentially unchanged binding characteristics. The suitability of ANA-CCK-8s for the purification of anti-CCK antibodies was also demonstrated. The approach may prove to be generally useful in the isolation of labile proteins in an intact form.