The article describes the development of an enzyme-immunoassay using antigen-enzyme conjugates, specifically human chorionic gonadotrophin (HCG) conjugated with horse radish peroxidase (HRP). The method involves preparing HCG-HRP conjugates through chemical coupling and purification by density gradient centrifugation. The conjugates are then used in immunoassays to detect HCG and antibodies against HCG. The study compares two immunoassay methods: the solid phase (SP) method and the double antibody solid phase (DASP) method. The DASP method showed higher sensitivity and better duplicate determinations compared to the SP method. The enzyme-immunoassay was tested on urine samples from pregnant and non-pregnant women, showing a good correlation with the haemagglutination inhibition assay. The sensitivity of the enzyme-immunoassay was comparable to that of haemagglutination inhibition or complement fixation assays, but the DASP method was 10-20 times more sensitive than the SP method. The study highlights the potential of enzyme-immunoassays for clinical applications, particularly in the detection of HCG in urine samples. The results demonstrate that the enzyme-immunoassay is a viable alternative to radio-immunoassays, offering similar sensitivity and accuracy. The study also discusses the importance of separating antibody-bound and free conjugates in enzyme-immunoassays, which is as critical as in radio-immunoassays. The method described is a significant advancement in immunoassay technology, providing a reliable and sensitive means of detecting HCG in clinical settings.The article describes the development of an enzyme-immunoassay using antigen-enzyme conjugates, specifically human chorionic gonadotrophin (HCG) conjugated with horse radish peroxidase (HRP). The method involves preparing HCG-HRP conjugates through chemical coupling and purification by density gradient centrifugation. The conjugates are then used in immunoassays to detect HCG and antibodies against HCG. The study compares two immunoassay methods: the solid phase (SP) method and the double antibody solid phase (DASP) method. The DASP method showed higher sensitivity and better duplicate determinations compared to the SP method. The enzyme-immunoassay was tested on urine samples from pregnant and non-pregnant women, showing a good correlation with the haemagglutination inhibition assay. The sensitivity of the enzyme-immunoassay was comparable to that of haemagglutination inhibition or complement fixation assays, but the DASP method was 10-20 times more sensitive than the SP method. The study highlights the potential of enzyme-immunoassays for clinical applications, particularly in the detection of HCG in urine samples. The results demonstrate that the enzyme-immunoassay is a viable alternative to radio-immunoassays, offering similar sensitivity and accuracy. The study also discusses the importance of separating antibody-bound and free conjugates in enzyme-immunoassays, which is as critical as in radio-immunoassays. The method described is a significant advancement in immunoassay technology, providing a reliable and sensitive means of detecting HCG in clinical settings.