Identification of poliovirions and subviral particles by capillary electrophoresis.
Poliovirions, purified from infected cell extracts with anion-exchange chromatography, can be analyzed and identified by CE in untreated fused silica capillaries using UV detection. Other subviral particles can be eluted as well from the same infected cell extract using a higher salt concentration buffer on the ion-exchange chromatography. Virions can be identified because of their conversion into empty capsids upon heating at 56°C. As a result of heating, the viral genome is released from the capsid. Here, we show that during this incubation some intermediate particles were found. The latter were identified by enzymatic peak shift analysis. The high salt concentration eluate subviral particles were analyzed with preincubation affinity CE together with their sensitivity for RNase and proteinase K treatment. Electropherograms of the higher salt concentration eluate display a mixture of at least four different subviral particles. One particle proved to have an [N1, H] antigenicity and was resistant to RNase and proteinase K digestion. The remaining particles were all sensitive to proteinase K treatment. This CE method proved to be valuable in the detection, identification and analysis of poliovirions and poliovirus particles offering an alternative powerful, cheap, fast and easy analysis method.
Department of Pharmaceutical Biotechnology and Molecular Biology, Center for Pharmaceutical Research, Vrije Universiteit Brussel, Brussels, Belgium. hhalewyc@vub.ac.be
Hadewych
H
Halewyck
Iuliana
I
Oita
Bert
B
Thys
Bieke
B
Dejaegher
Yvan
Y
Vander Heyden
Bart
B
Rombaut
D000911:Antibodies, Monoclonal / Q000737:chemistry; D000914:Antibodies, Viral / Q000737:chemistry; D000956:Antigens, Viral / Q000737:chemistry / Q000302:isolation & purification; D036022:Capsid Proteins / Q000737:chemistry; D002852:Chromatography, Ion Exchange; D019075:Electrophoresis, Capillary / Q000379:methods*; D019286:Endopeptidase K / Q000378:metabolism; D016679:Genome, Viral; D006358:Hot Temperature; D017955:Poliovirus / Q000737:chemistry*; D012260:Ribonucleases / Q000378:metabolism; D014771:Virion / Q000737:chemistry / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; D012260:Ribonucleases; D019286:Endopeptidase K