[A high-capacity affinity gel for the purification of the testicular lutropin receptor]

The lutropin (LH) receptor from porcine testes was solubilized with 1% Triton X-100. The final solution was centrifuged at 100,000 X g and the supernatant was concentrated on a PM30 Amicon membrane. The concentrate was then specifically retained on an affinity gel with human choriogonadotropin (hCG) covalently grafted by its sugar moiety. The agarose affinity gel bore adipic acid hydrazide spacer arms (6 carbon length). Prior to coupling, the subunits of hCG were crosslinked with 1-ethyl-3-(dimethylamino propyl) carbodiimide and the sialic acid residues were subjected to periodate oxidation to yield aldehyde groups which could react with the hydrazides of the gel. 90% of the receptor contained in the solution incubated with this gel was retained and eluted at pH 3.2 with a purification factor greater than 700, versus a 26% yield and a purification factor of 40 with an affinity gel bearing hCG linked through its lysine epsilon-amines.