RADIOIMMUNOASSAY FOR HUMAN LIPOPROTEIN LIPASE

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The purpose of this research project is to develop and validate a precise and sensitive non-competitive solid phase radioimmunoassay for lipoprotein lipase (LPL) for human pre- and post-heparin plasma. The method has already been developed in our laboratory for avian LPL. In order to set up a similar assay for human LPL, it will be necessary to purify bovine milk LPL to homogeneity. Antisera against this enzyme will be produced in goats and immunoglobulins purified on LPL-Sepharose affinity columns. The immunoglobulins will be coupled to hydrophilic polyacrylamide beads. These immunobeads will be employed in conjunction with 125I immunoglobulins in a direct sandwich-type radioimmunoassay. The precision and accuracy of the assay will be evaluated, as well as possible interference by plasma proteins and exogenous chemicals such as heparin. LPL activities in normal and hyperlipemic pre- and post-heparin human plasma will be determined. A similar radioimmunoassay will be developed employing human LPL and anti-LPL immunoglobulins. The physical and chemical properties of highly purified human LPL will be compared to those of liver lipase. The hypothesis that successive binding and release of triglyceride-rich lipoproteins with the vascular endothelium leads to a gradual enrichment of the lipoprotein with LPL proteins will be tested. The molar ratios of LPL and VLDL subfractions resolved by rate zonal ultracentrifugation will also be determined.