Abstract
A quantitative reversed-phase high-performance liquid chromatography (RP-HPLC) method has been developed to detect the L1 subunit protein from virus-like particles (VLPs) of human papillomavirus (HPV). The method utilizes heat treatment with a buffer consisting of 50 m M Tris, pH 8.0 containing 8 M guanidine–HCl and 10% 2-mercaptoethanol to dissociate the VLPs into monomeric L1. Following dissociation, the sample is injected onto a C 4 or C 8 column. The L1 protein is eluted as a single, clearly resolved peak. Elution conditions have been optimized to enhance the separation of L1 from other contaminants. Based on spike recovery studies and sodium dodecyl sulfate–polyacrylamide gel electrophoretic analysis this method is suitable for quantitation of various partially purified in-process samples and can be used to facilitate purification process development.
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