Production and characterization of a monoclonal antibody for sweat-specific protein and its application for sweat...

Identification of body fluids is a common task in medico-legal practice, but specific markers for sweat have not been identified to date. To develop a method for identification of sweat, we identified a sweat-specific protein and produced monoclonal antibodies by immunizing mice with sweat proteins fractionated by anion-exchange chromatography. Among many sweat-reactive monoclonal antibodies obtained, one monoclonal antibody (G-81) was selected because of its unique specificity. G-81 reacted to sweat but not to other body fluids (e.g. serum, saliva, semen, milk, urine and tears) in ELISA. G-81 specifically stained the eccrine sweat gland and did not stain any other tissue including the apocrine sweat gland. In western blotting, G-81 reacted strongly to a 7 kDa band and faintly to 20, 27 and 33 kDa bands of sweat protein. The N-terminal amino acid sequence (18 amino acids) of G-81-reactive peptides was determined, and an identical sequence was found in an antibiotic peptide dermcidin (110 amino acids) reported recently, suggesting that G-81 recognized a fragment of dermcidin. The G-81-reactive peptide could be detected in 8,192-fold dilutions of sweat by ELISA and could be detected in 200-fold diluted sweat samples ( n=26) independent of the protein concentration. The G-81-reactive peptide was very stable and was able to detect sweat stains left for at least 11 weeks at room temperature without substantial loss of reactivity. These facts suggest that G-81 is a very useful tool for sweat identification in medico-legal practice.