A sandwich enzyme-linked immunosorbent assay for ABO blood typing of semenby using anti-p 84 monoclonal antibody as a marker of blood group substance in semen

A blood group substance (BGS), a protein with ABH antigenic activity, was i solated from human seminal plasma and designated as p 84 (Sato, 1995) (1). We have developed a method for determining the ABO blood type of semen by p erforming a sandwich enzyme-linked immunosorbent assay (ELISA) in which p 8 4 is captured with an anti-p 84 monoclonal antibody, and evaluated the spec ificity and sensitivity of this method. Although BGS activity was detected in semen sensitively by this method, it was not detected in saliva, urine, breast milk, blood or vaginal secretions. Since the concentration of p 84 i n semen was independent of the secretion status, the status can be determin ed as non-secretor when p 84 but not BGS activity was detected. To determin e the stability of BGS activity on p 84, dried stains of semen on filter pa per were kept at 4, 26, and 37 degrees C for 8 months, 2 years and 1 month, respectively, and their BGS activities were examined. After 8 months at 4 degrees C, over 60% of the original BGS activity was recovered from the sta in. The activity could be detected even from a square as small as 0.25 by 0 .25 cm. After 1 month at 37 degrees C and 2 years at 26 degrees C, 31 and 2 0% of the BGS activity, respectively, still remained. It could be detected from the pieces of 1.0 by 1.0 cm and 0.5 by 0.5 cm squares, kept for 1 mont h at 37 degrees C and 2 years at 26 degrees C, respectively. Finally, semen was mixed with saliva or blood at varying volumetric ratios and used for t he sources of dried stains. The BGS activity of p 84 could be detected in t he stains until the ratio between semen and saliva or blood reached 1:4. We conclude that this sandwich ELISA offers a more sensitive and specific met hod for determining the ABO blood type of semen samples obtained from sexua l assault victims than existing methods, such as the conventional absorptio n-elution and classical hemagglutination-inhibition tests.