Detection and semi-quantitative determination of low abundance GFAP mRNA in mouse brain by capillary electrophoresis coupled with laser-induced fluorescence.
The sensitivity of capillary electrophoresis coupled with laser-induced fluorescence (CE-LIF) was compared with conventional agarose gel electrophoresis-ethidium bromide-UV method (AE-EUV) for detection and semi-quantitative determination of GFAP mRNA in mouse brain. GFAP expression was induced by the neurotoxin MPTP in C57BL mice. Serially diluted RNA samples (0.0003, 0.003, 0.03, 0.3, and 3 microg total RNA) were subjected to RT-PCR and analyzed by both procedures. The integrated pixel density (AE-EUV) and peak area (CE-LIF) were directly proportional to the amount of RNA. However, the observed high sensitivity of CE-LIF suggests its potential application for detection and semi-quantitative determination of low-abundance mRNA transcripts.
Khan
Haseeb Ahmad
HA
Department of Biochemistry, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia. khan_haseeb@yahoo.com
D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; D000818:Animals; D001253:Astrocytes / Q000737:chemistry* / Q000187:drug effects; D001923:Brain Chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D005453:Fluorescence; D005904:Glial Fibrillary Acidic Protein / Q000235:genetics*; D005911:Gliosis / Q000139:chemically induced / Q000175:diagnosis*; D007834:Lasers; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis*
D005904:Glial Fibrillary Acidic Protein; D012333:RNA, Messenger; D015632:1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine