Key result
LNA flow-FISH tracks early Sindbis virus RNA with comparable sensitivity to EGFP constructs.
Why the study?
A straightforward and broadly applicable method to monitor viral RNA in infected cells was needed to complement existing quantification techniques.
LNA flow-FISH is a sensitive and straightforward method for monitoring viral RNA replication and the effects of antiviral medication in infected cells.
May aid preclinical antiviral screening; leaves open clinical translation and patient outcome impact.
We previously showed the feasibility of using locked nucleic acid (LNA) for flow cytometric-fluorescence in situ hybridization (LNA flow-FISH) detection of a target cellular mRNA. Here we demonstrate how the method can be used to monitor viral RNA in infected cells. We compared the results of the LNA flow-FISH with other methods of quantifying virus replication, including the use of an enhanced green fluorescent protein (EGFP) viral construct and quantitative reverse-transcription polymerase chain reaction. We found that an LNA probe complementary to Sindbis virus RNA is able to track the increase in viral RNA over time in early infection. In addition, this method is comparable to the EGFP construct in sensitivity, with both peaking around 3 h and at the same level of infected cells. Finally, we observed that the LNA flow-FISH method responds to the decrease in levels of viral RNA caused by antiviral medication. This technique represents a straightforward way to monitor viral infection in cells and is easily applicable to any virus.
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Robertson et al. (2010) studied Viral infection. LNA flow-FISH vs. EGFP viral construct and quantitative RT-PCR was evaluated on Detection and quantification of viral RNA. LNA flow-FISH successfully tracked the increase in Sindbis virus RNA during early infection, showing comparable sensitivity to an EGFP construct with both peaking around 3 hours.
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