Diagnostic Detection of Herpes Simplex and Hepatitis C Viral Amplicons by Capillary Electrophoresis: Comparison With Southern Blot Detection.
Background: As a result of the large number of DNA-based clinical assays, there is intense interest in making polymerase chain reaction (PCR)-amplified DNA product analysis faster, more cost-effective, and more automated. Methods and Results: In this study, an evaluation of the use of capillary gel electrophoresis with laser-induced fluorescence detection is described as a means of analyzing postamplification PCR products from clinical specimens. Sixty-six herpes simplex virus and 152 hepatitis C virus amplicons were analyzed after PCR and reverse transcription PCR, respectively. It is shown that the use of a physical gel buffer system in a short capillary in conjunction with laser-induced fluorescence detection allows for sensitive detection of herpes simplex virus- and hepatitis C virus-specific DNA fragments in an expedient manner. Interinstrument and intercapillary reproducibility of the migration time was evaluated and found to be excellent. The advantages and disadvantages over agarose gel electorphoresis-Southern blot analysis are summarized. Conclusions: The advantages offered by capillary gel electrophoresis with laser-induced fluorescent detection including rapid and sensitive analysis, ease of setup, reduced cost, and possibility for automation, make this procedure a viable alternative to more labor-intensive agarose gel electrophoresis-Southern blot analysis as molecular diagnostic methodology.
Pancholi
P
Department of Laboratory Medicine and Pathology, Mayo Foundation/Mayo Clinic, Rochester, Minnesota, USA
Oda
RP
Mitchell
PS
Landers
JP
Persing
DH