Rapid and Easy Separation of the Subunits of Bovine and Human Glycoprotein Hormones by Use of High Performance Liquid Chromatography*

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Abstract
A single method of reverse phase high performance liquid chromatography is used to separate the subunits of human and bovine glycoprotein hormones. This rapid and easy method is applicable for the separation and detection of subunits from as little as 10 .mu.g hormone or the isolation of subunits from as much as 100 mg hormone. Separation is achieved by chromatography on a Vydac 218TP1010 column with a linear (60 min) gradient of 0.1 M sodium phosphate, pH 6.8, plus 1 mM sodium azide to a solvent containing 50% acetonitrile and 50% 0.1 M sodium phosphate, pH 6.8, plus 1 mM sodium azide. Although in some cases the interaction between the hydrophobic support and the hormone is sufficient for dissociation, preincubation of the hormone with guanidine hydrochloride ensures optimum dissociation and improves resolution of the subunits. The subunits isolated by high performance liquid chromatography are functional in that they will reassociate with their counterpart subunits.