SpermPaint Optimization and Validation

• NCJ Number: 220289
• Author(s): Dr. John Herr
• Date Published: March 2007
• Length: 20 pages
• Grant Number(s): 2000-IJ-CX-K013
• Agencies: NIJ-Sponsored
• Publication Type: Report (Study/Research)
• Award: SPERM CELL SELECTION SYSTEM FOR FORENSIC DNA OF THE MALE COMPONENT

Annotation

This report presents the methodology and findings of the SpermPaint Project, whose goal was to develop monoclonal antibody probes that would facilitate the rapid identification of sperm when the number of sperm are low in samples collected with sexual-assault swabs.

Abstract

The study found that the fluorescent signal from both monoclonal antibodies (equatorial segment protein [ESP] and calcium binding tyrosine phosphorylation regulated protein [CABYR]) was clear and bright with no cross reactivity to other tissue. SpermPaint demonstrated that it is effective in identifying sperm in postcoital samples that were stored at 4 degrees C for 2 years. The target proteins were stable in samples collected at 1, 24, and 72 hours. It positively identified questionable shapes, sperm adherent to epithelial cells, and sperm hidden within cellular debris. Twenty isolated sperm were sufficient to amplify a 350 based pair PCR product in the majority of experiments. In contrast, 10 sperm yielded results less than 50 percent of the time. These preliminary results indicate that the collection of 20 sperm or sperm flagella provides a good starting point for amplifying mtDNA in the proposed experiments. It is anticipated that more sperm will be identified with SpermPaint compared with current stain methods. The current formulation of SpermPaint is a mixture of fluorescent dye conjugated monoclonal antibodies that allows definitive identification of human sperm by using fluorescence microscopy. The current formulation of SpermPaint contains a monoclonal antibody specific for the sperm head antigen ESP and a monoclonal antibody specific for the sperm flagellar antigen CABYR. Together, this mixture of monoclonal antibodies stains the sperm head with a characteristic band pattern through its central region corresponding to the equatorial segment and stains the longest domain of the sperm tail corresponding to the principal segment. 11 figures, 1 table, 17 publications listed, 12 references, and appended SpermPaint protocol