Possible synthesis of steroid-protein for immunization with a fixed narrow range of the hapten-protein ratio

A comparison of two methods for synthesis of a steroid-protein conjugate (progesterone-11 alpha-hemisuccinate-BSA) has been made. Under the conditions of the method described, using tetrahydrofurane as a medium for the coupling reaction, stable intermediate products were obtained. The resulting conjugate had a narrow range of the hapten-protein ratio (18-20 steroid molecules per molecule of BSA), very good solubility in water and almost no unreacted BSA in the sample. Antisera raised against this conjugate, applied in low dose (50 micrograms), had quite satisfactory characteristics concerning their titre and specificity. The tests for recovery, sensitivity and the range of measurement established the possibility of using such antisera for the radioimmunoassay of progesterone.