ISSN: 0022-1198
CODEN: JFSCA
Page Count: 6

An Alternative, Effective Substrate for Erythrocyte Acid Phosphatase Phenotype Determinations
Gambel, AM
Research chemist and scientist, Forensic Science Research and Training Center, Laboratory Division, FBI Academy, VA

Budowle, B
Research chemist and scientist, Forensic Science Research and Training Center, Laboratory Division, FBI Academy, VA

(Received 27 July 1987; accepted 13 November 1987)

Abstract

A method is described for obtaining nondiffusing, nonfading fluorogenic zymograms for erythrocyte acid phosphatase variants separated by isoelectric focusing. The synthetic substrate 4-trifluoromethylcoumarin phosphate was impregnated into cellulose diacetate membranes and air-dried overnight. After isoelectric focusing. the substrate overlay membrane was rehydrated in 0.05M citrate buffer, pH 4.0, lightly blotted, and overlayed on the gel. A 5- to 10-min incubation at 37°C produced intensely fluorescing, light-blue bands on the diacetate membrane. Interaction of the trifluoromethyl group on the substrate with hydrophobic regions of the diacetate membrane impeded diffusion, yielding a permanent zymogram. There were no discrepancies in phenotype determinations using this method when compared with the 4-methylumbelliferyl phosphate assay approach. Further, an increased number of conclusive calls were obtained (91.8 versus 79.5% and 54.1 versus 34.9%) with this new assay when compared with the 4-methylumbelliferyl phosphate substrate on known liquid bloods and questioned dried bloodstains, respectively.

Keywords:
forensic science, phosphatases, fluorogenic substrates, isoelectric focusing, cellulose diacetate membranes, erythrocyte acid phosphatase, forensic serology, bloodstains

Paper ID: JFS12514J
DOI: 10.1520/JFS12514J
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