An enzyme immunoassay for human serum paraoxonase 1 (PON 1)
Borriello, Erica A.
MetadataShow full item record
Human Serum Paraoxonase 1 (PON 1) plays a dual physiological role. Low PON status levels are an emerging risk factor for coronary vascular disease (CVD) where it is has been to shown to be responsible for HDL's (high-density lipoproteins) ability to protect LDL (low-density lipoproteins) from oxidative modification. PON is also a critical player in the metabolism of organophosphate insecticides and other environmental intoxicants. The analysis of PON1 requires the measurement of three factors, which directly and significantly impact the role of this enzyme in humans: (1) polymorphism classification, (2) enzymatic activity, and (3) enzyme concentration. The combination of these three factors referred is referred to as 'PON 1 Status'. Determination of an individual's PON 1 status is necessary to accurately classify an individual in terms of relative risk for CVD as well as susceptibility to environmental poisoning. Previous work has produced ways to measure PON 1 activity and determine molecular polymorphisms. To date, no commercial assay has been developed to measure PON 1 concentration. There is consequently a need to develop a PON 1 Enzyme Linked Immunosorbant Assay (ELISA) to facilitate the quantification of the enzyme in human serum. The development of a functional ELISA will enable researchers to determine overall PON 1 status of individuals, measuring concentration in addition to genotype. The specific aims of this project are to: (1) Culture, purify and characterize a previously developed mouse-anti human PON 1 monoclonal antibody. (2) Prepare human PON 1 protein standard by isolation and purification assays from large units of human plasma. (3) Generate, purify and characterize a polyclonal goat anti-human antibody. (4) Design a PON 1 ELISA using the standards, monoclonal antibody and polyclonal antibody to produce a functional, sensitive and rapid assay to measure human serum PON 1. The results of this research will provide the necessary PON 1 concentration assay to complement the phenotype and activity assays already established. These 3 assays will provide the comprehensive analytical tools necessary to more thoroughly investigate the role of PON1 in human health and disease states.