SYNERGIES BETWEEN MICROPREPARATIVE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AND AN INSTRUMENTAL OPTICAL BIOSENSOR

The recent development of an automated surface plasmon resonance techn ology for the measurement of biomolecular interactions (Pharmacia BIAc ore) has provided new opportunities for the detection and analysis of protein-protein interactions. In the BIAcore, detection is based on ch anges in surface plasmon resonance which are monitored optically. Chan ges in surface plasmon resonance correspond to changes in surface conc entration of macromolecules and can be monitored in real time. We have found that the detection sensitivity obtainable with this technology (ng/ml concentrations of specific ligands are readily detectable for m any applications) is complementary ''in a bidirectional manner'' to mi cropreparative HPLC. Thus micropreparative HPLC may be used to purify and characterise reagents for the biosensor, whilst the biosensor may be used to define chromatographic parameters such as elution condition s for affinity chromatography or serve as an affinity detector for fra ctions obtained during chromatographic purification. Examples of such applications, including the potential of the biosensor to search for a nd monitor the purification of unknown ligands for which the target mo lecule has been identified, are shown. In particular, the use of the b iosensor to monitor the purification of soluble epidermal growth facto r receptor from A431 cell conditioned media is demonstrated.