A Review of “Viral Quantitative Capillary Electrophoresis for Counting Intact Viruses”
Mironov, G.G.; Chechik, A.V.; Ozer, R.; Bell, J.C.; Berezovski, M.V. Analytical Chemistry.
2011, 83, 5431-5435
Kirsten Berube
March 5, 2012
This study attempts to establish a method for virus quantification based on capillary
electrophoresis instrumentation. Quantification of viruses involves counting the number of
viruses in a given volume to determine the viral concentration. It is very useful in research for
possible cancer therapies involving oncolytic viruses. The method introduced in this article is
named viral quantitative capillary electrophoresis (viral qCE) and is a separation technique that
distinguishes between intact virus particles and residual DNA using capillary electrophoresis
with laser induced fluorescence detection. They used the JX-594 strain of vaccinia virus (VV),
which is a member of the poxvirus family and is designed to attack cancer. A mixture of virus
particles and free DNA was separated by capillary electrophoresis. An intercalating dye was used
to detect encapsulated DNA and free DNA and an anti-VV antibody labeled virus peaks. Two
zones emerged. There was a fast-moving zone made of intact particles presented as a group of
narrow peaks, and there was a slow-moving zone made of free DNA observed as a wide peak.
Virus lysis by heat and proteases caused the fast-moving zone to disappear and the slow-moving
zone of free DNA to intensify. The contamination level of VV was calculated using the free
DNA peaks before and after lysis. The concentration of viral DNA was determined by using a
calibration curve created from serial dilutions of phage DNA. Using a formula, the concentration
of intact virus particles was calculated. To confirm the quantitation of intact viral particles, a
technique involving laser-illuminated microscopy called nanoparticle tracking analysis (NTA)
was also used. Three samples of VV were studied including 2 month old VV, VV that underwent
20 freeze/thaw cycles, and freshly prepared virus. Lysis of the virus was studied by different
methods including temperature, sonication, SDS, and proteinase K. Overall, they found three
advantages of viral qCE: it works in a wider range of virus concentrations, its sample depletion is
very low, and its ability to calculate the level of virus contamination by denatured viral DNA or
residual cellular DNA.
Electrophoresis seems to be a useful method for viral quantitation. It makes sense that
intact viral DNA and DNA particles could easily be separated on a capillary by an electric field
since it involves separating charged particles of different sizes. The researchers created a
calibration curve which is an effective way to determine an unknown measurement from a
known measurement based on standards. They also backed up their calculated results with a
technique currently used for quantitation of viruses that is based on laser-illuminated microscopy
where each particle’s movement and speed is tracked, allowing it to determine the concentration
and size of particles. The study mentioned a number of techniques used for viral quantitation
such as ELISA and qPCR. It would have been more informative if they included comparisons
between these techniques and capillary electrophoresis results since those techniques are
common. The study included three different VV samples and different virus lysis methods.
Having too many variables complicates the study and can affect results. Some of these variables
could have been regarded in separate studies to reduce the chance of error in this study.
Overall, it seems that a new, affective method for viral quantitation has been performed.
The science of the separation and calculations make sense and advantages of capillary
electrophoresis have been found. The study could have had fewer variables and additional
studies including these variables could be carried out.