Abstract
Fundamental studies were made on hemagglutination inhibition reaction (HAIR) method with anti-estriol 16-glucuronide (E3-16-G) serum for a rapid and simplified determination of estrogen in pregnancy urine. The sheep erythrocytes fixed with formalin and treated with tannic acid were sensitized with E3-16-G-rabbit serum albumin (RSA) conjugates. An antiserum to E3-16-G-bovine serum albumin (BSA) was absorbed with both BSA and erythrocytes fixed with formalin. The degree of binding of E3-16-G with RSA and the amount of E3-16-G-RSA sensitized onto erythrocytes affected the easiness of adjusting sensitivity and the clearcut patterns of HAIR formed at the bottom of the test tubes. A HAIR reagent having sensitivity of 0.2 μg/ml (E3-16-G equivalent in E3) could be prepared by sensitizing 8% erythrocytes suspension with a 0.05 to 0.08% of E3-16-G-RSA in which 20 to 30 mol of E3-16-G being bound per 1 mol RSA, and finaly the sensitivity was adjusted by diluting an antiserum. Cross-reactivity of the HAIR method with estrogens was almost similar to that of radioimmunoassay (RIA) and the reactions were found with estrogens other than conjugated at the C-3 position i.e., E3-16-G, C-17 conjugated estrogens, free estrogens. Glucose, NaCl, serum protein, urinary protein and urine pH did not affect the results. E3-16-G added to pregnancy urine was recovered in satisfactory yield by considering the accuracy of the method. Significant correlation (y=0.78x+0.97, r=0.949) was noted between HAIR and RIA on the measured estrogen values of the same urines from pregnant women. These results indicate that our HAIR method, semi-quantitative determination of urinary estrogen, is a useful tool for feto-placental function in clinical routine.
