A new assay for β-thromboglobulin in urine

Abstract

Measurements of β-thromboglobulin (βTG) excretion in urine may be of value for “field” studies and due to problems with sampling artifacts for BTG in plasma. Previous studies have used a radioimmunoassay designed for plasma without characterizing the “BTG” immunoreactivity in urine. We describe modifications of the assay which increase its sensitivity and a sample work-up procedure using Sephadex G-25M columns separating high molecular weight (HMW) components (presumably intact βTG) from low molecular weight (LMW) immunoreactivity (i.e. βTG fragments and/or non-specific interferences). The sensitivity of the assay (with 2.5 ml sample) is <12 pg/ml HMW BTG. Inter-and intraassay coefficients of variation were 7–10%. Only 33 (range 5–75)% of BTG immunoreactivity in urine represented HMW βTG. LMW immunoreactivity may be related to salt and other non-specific influences in the sample. Recoveries of βTG were quite variable (9–100%) in unextracted urines, but high and reproducible (80±2%) in the HMW fraction. Thus, nonspecific interferences with BTG measurements in certain urines are overcome by the separation step. Using Sephadex fractionation βTG immunoreactivities in night urines (n=15) were: 20±3 pg/ml in the HMW fraction, 70±8 pg/ml in the LMW fraction, and 85± 10 pg/ml by direct assay. HMW βTG increased in daytime samples (to 30±5 pg/ml; p<0.01), but no diurnal variation was seen in the LMW fraction or with the direct assay. Thus, selective analysis of HMW βTG in urine circumvents problems with nonspecific immunoreactivity and apparent interferences with measurements of intact βTG. The present more selective assay for HMW immunoreactivity increases the possibility of detecting physiological changes in βTG release in vivo by urinary measurements.