A rapid versatile microassay for cellular retinol-binding protein using Lipidex-1000 microcolumns.

A new, rapid and versatile microassay for cellular retinol-binding protein has been developed based on separation of bound and free ligand by means of Lipidex-1000, a hydrophobic Sephadex derivative. This requires quantitative manipulation of retinol in aqueous solution. The tendency of retinol to adhere to glass and plastic surfaces was overcome by addition of the detergent Ammonyx LO, which yields a micellar dispersion.

Phosphorylation reactions in bovine rod outer segments studied by 32P-labelling of intact retina.

The protein phosphorylation pattern in the intact bovine retina has been investigated by labelling with 32P-phosphate under incubation conditions that preserve the electrical photoresponse of the photoreceptor cells. The phosphorylation of rod outer segment proteins was analysed after isolation of outer segments from the labelled retina. The global influence of light, Ca2+ and the phosphodiesterase inhibitor, isobutylmethylxanthine, on protein phosphorylation in rod outer segments was analysed.

A new isolation procedure for retinal pigment epithelium.

A new isolation procedure for bovine retinal pigment epithelial cells has been developed. It is based on perfusion of the whole bovine eye via the central ophthalmic artery with a cold, buffered isotonic salt solution free of divalent cations for 15 min. The perfusion both weakens the association of the pigment epithelial cells with Bruch's membrane and the adhesion between retina and pigment epithelium.

Rod outer segment membranes: good acceptors for retinoids?

The exchange of all-trans retinoids (retinal, retinol, retinylpalmitate) between PC-vesicles, PC-vesicles and liver microsomes or PC-vesicles and rod outer segment membranes is investigated using 11,12(3)H labeled compounds. In the first two systems, retinal and retinol exchange rapidly, retinyl acetate slowly and retinyl palmitate not at all. Rod outer segment membranes however take up relatively small amounts of retinoids (retinylpalmitate less than retinol less than retinal) and rapidly lose 60-90% of their label in the presence of PC-vesicles.