Quantitative Sandwich Enzyme-linked Immunosorbent Assay for Human Secretory Prostaglandin D Synthase (.BETA.-Trace).

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Glutathione-independent prostaglandin D synthase is responsible for the biosynthesis of prostaglandin D2, an endogenous sleep-promoting substance, in the central nervous system of various mammals including humans. This enzyme is localized in the choroid plexus, leptomeninges, and oligodendrocytes of the central nervous system and is secreted into the cerebrospinal fluid as β-trace protein, a major constituent of human cerebrospinal fluid. Two monoclonal antibodies against human β-trace (1B7 and 10A3) were prepared by immunization of BALB/c mice with the recombinant protein expressed in Escherichia coli. Western blot analysis with human cerebrospinal fluid revealed that both 1B7 and 10A3 antibodies were immunoreactive toward a single protein at the same position as that of the purified β-trace (Mr=27, 000). A quantitative sandwich enzyme-linked immunosorbent assay was constructed with these two monoclonal antibodies. The assay system showed a linearity in the range from 0.06 to 4.00ng β-trace/well (100μl). The β-trace concentration was determined by the immunoassay to be 11.30±2.54μg/ml in human cerebrospinal fluid, 1.25±0.37μg/ml in the urine, and 0.27±0.01μg/ml in the serum.
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