Detection of dermcidin for sweat identification by real-time RT-PCR and ELISA.
We evaluated the performance of real-time RT-PCR and ELISA assays for detection of dermcidin (DCD) in sweat and body-fluid stains. DCD, a small antibiotic peptide secreted into human sweat, was detected by real-time RT-PCR in 7-day-old stains containing as small as 10 microL of sweat, and the assay showed high specificity when testing 7-day-old stains containing 30 microL of other body-fluid. ELISA using anti-human dermcidin mouse monoclonal antibody detected DCD sweat diluted up to approximately 10,000-fold and could specifically detect DCD in 10 microL of body-fluid stains. The performance of the two assays was tested during winter on samples that simulated forensic case samples: an undershirt and a sock worn for 20 h, a handkerchief used to wipe the brow several times within 12h, a cap and a cotton glove worn for 4h, and a white robe worn at intervals for 2 years. The result showed that the former assay detected DCD in all sites of the undershirt examined (armpit, back, and breast), and the latter gave a relatively high OD value in the armpit among the three sites. For the socks, although the latter assay gave very high OD values in both the center and toe of the foot sole, the former could not detect DCD in both of them. These results indicate that highly damp conditions, such as inside a shoe, might promote the degradation of mRNA in samples such as socks. In the other case samples, sweat was adequately detected by both assays. This study is the first demonstration of the use of real-time RT-PCR to sensitively identify sweat among body-fluid stains, and it confirmed that dermcidin was an excellent marker for sweat identification. In addition, the usefulness of ELISA was also verified. Positive sweat identification using these assays is expected to assist forensic practice.
Sakurada
Koichi
K
National Research Institute of Police Science, Kashiwa, Chiba, Japan. sakurada@nrips.go.jp
Akutsu
Tomoko
T
Fukushima
Hisayo
H
Watanabe
Ken
K
Yoshino
Mineo
M
D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D001769:Blood / Q000378:metabolism; D002582:Cervix Mucus / Q000378:metabolism; D003020:Clothing; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005554:Forensic Medicine; D006801:Humans; D008297:Male; D008875:Middle Aged; D010455:Peptides / Q000235:genetics* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012313:RNA / Q000378:metabolism; D012463:Saliva / Q000378:metabolism; D012661:Semen / Q000378:metabolism; D012680:Sensitivity and Specificity; D013542:Sweat / Q000378:metabolism*; D014556:Urine
D015415:Biomarkers; D010455:Peptides; C442243:dermcidin; D012313:RNA