Quantitation of RNA tumor viruses by spectroscopy of density gradient gels.
We have developed a system for virus particle quantitation based on the measurement of the optical absorbance of stained viruses which first have been banded at their buoyant density in an equilibrum 24 to 53% (wt/wt) sucrose density gradient, then fixed in position in the gradient by photopolymerizing an acrylamide-riboflavin mixture in the sucrose, and finally stained and destained. Using plasma from mice infected with leukemia virus (Rauscher) or chickens infected with avian myeloblastosis virus (BAI strain) or suitable controls, we have shown that this technique specifically detects RNA tumor viruses. By using virus stock solutions for which the absolute concentrations were determined by laser beat frequency spectroscopy, we have calibrated the absorbance of the viral bands in terms of virus particle concentration. Using 0.8-ml gradients gels (4 by 45 mm) we can detect as low as 2 x 10(7) viral particles with Coomassie blue staining and 6 x 10(6) viral particles with a more sensitive staining procedure using amido black.
D000818:Animals; D009189:Avian Myeloblastosis Virus / Q000302:isolation & purification; D002499:Centrifugation, Density Gradient; D002645:Chickens; D007834:Lasers; D009052:Leukemia Virus, Murine / Q000302:isolation & purification; D051379:Mice; D009858:Oncogenic Viruses / Q000302:isolation & purification*; D012328:RNA Viruses / Q000302:isolation & purification*; D013057:Spectrum Analysis / Q000379:methods*; D013194:Staining and Labeling