Identification of poliovirions and subviral particles by capillary electrophoresis

Abstract

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 561C. 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.

Poliovirus separation from cell extracts using capillary electrophoresis: Potential use in vaccine production and control?

“…Capillary electrophoresis was used to quantitate the poliovirus particles (Oita et al, 2011), and to distinguish the D-antigen from the heat denaturated poliovirus, non-exposing the D-antigen (Halewyck et al, 2010). Additionally, Okun et al (2000) demonstrated that another picornavirus, the human rhinovirus, was still viable after a similar capillary electrophoresis procedure.…”

Rational use of stacking principles for signal enhancement in capillary electrophoretic separations of poliovirus samples

“…Compared to real life virological samples, this is still a high concentration, and it was expected that this method will not be useful to study PV from cell extracts, where much lower concentrations may occur. Also, when highly concentrated PV samples are degraded upon heating, or viral-assembly particles are injected, the low level of the signal will make the investigation of those sub-viral particles extremely difficult [33]. Therefore a signal-increasing approach, also applicable to less pure PV samples, was necessary as an elegant solution to the problems mentioned above.…”

“…1A trace (a)). The use of SGPV samples for method development was necessary to simplify the peak identification, which is difficult in the absence of reference materials for PV [33]. However, for this method, the possible applications are limited to PV identification and quantification, after extensive purification of the original samples.…”

“…The earlier method failed to detect PV from the samples below 90 g/ml PV (data not shown). The proposed strategy also should allow parallel investigation of PV, subviral particles and different AEX fractions [33].…”

Recent developments in capillary and chip electrophoresis of bioparticles: Viruses, organelles, and cells

“…Composition of the BGE and parameters such as temperature, voltage, and injection conditions were varied (see Table 1 for the conditions used in this and in the following papers). The same analysis conditions, including SDS as additive, as in [8], were used in a subsequent paper [9], and subviral particles obtained by incubation at 561C were separated and identified. Furthermore, these authors achieved separation of the viral genomic RNA, empty capsids, and a not further characterized ''ribonucleoprotein.''…”