Chemiluminescent detection of DNA probes in forensic analysis.

The conditions for hybridization and detection of enzyme-labeled probes have been optimized in our laboratory for use with oligonucleotides coupled to alkaline phosphatase. We have examined several enzyme-linked probe which are complimentary to commonly used variable number of tandem repeats (VNTR) loci to determine the feasibility of using chemiluminescence for routine application in forensic DNA analysis. It was found that a chemiluminescent detection system employing an alkaline phosphatase activated dioxetane in the presence of chemiluminescent enhancers provides a high degree of sensitivity in hybridization protocols with a significant savings in overall filter processing time. The chemiluminescent system achieved equal or greater sensitivity that observed for 32P-labeled probes in much shorter development times. Furthermore, a new chemiluminescent substrate, Lumi-Phos Plus, has recently been investigated and found to further decrease the filter development time for forensic assays.