Virucide usage is extremely common, particularly within farming, veterinary, and hospital environments. Virucides prevent infection from occurring by inactivating viruses that are present on solid surfaces (such as countertops, door handles, or hands) or in liquid suspension. Traditionally, inactivation of viruses is evaluated by observing the development of CPE in permissible cellsin plaque assays. Viral CPE evaluation by plaque assay is subjective and there is no establish standard on scoring of CPEs. In contrast, the xCELLigence Real Time Cell Analysis system evaluates viral CPE in virucidal efficacy testing with precise measurement and extraordinary reproducibility.
Typical methods vs xCELLigence RTCA
|Plaque Assay||xCELLigence RTCA Viral Fitness Assay|
|Counting of number of plaques formed by a virus at varying dilutions to obtain a low multiplicity of infection (MOI). Concentration of virus (titer) is calculated as plaque forming units (PFU) per unit of volume.||Direct, real time, and quantitative kinetics for the entire virus life cycle. Cells are plated, infected with virus, and xCELLigence RTCA automatically monitors viral fitness in real time.|
|Viral plaque formation can take days to weeks to be detectable.||Quantitative monitoring of both fast (hours) and slow (days to weeks) CPE.|
|Plaque assay provides no information about the onset of CPE or the kinetics of virus-mediated cytotoxicity.||xCELLigence provides assessment and quantification of the full virus life cycle.|
|The manual counting of plaques by visual inspection can be highly subjective and inconsistent since plaque formation rates and sizes can vary dramatically.||Measurements are automatically recorded at a user defined frequency and are plotted by the xCELLigence software as Cell Index (CI). Accurate, precise, and highly reproducible method with less manual labor.|
Example Data: Evaluating the inactivation of Infectious Bursal Disease Virus (IBDV) Using a Virucide
Reprinted from the Journal of Virological Methods, volume 199, Ebersohn, K. et al. “An Improved Method for Determining Virucidal Efficacy of a Chemical Disinfectant Using an Electrical Impedance Assay,” pages 25–28. Copyright 2014, with permission from Elsevier.
Protecting Vero cells from IBDV using virucide. Vero cells infected with serial dilutions of IBDV that had not been pre-treated with virucide (A), or which had not been pre-treated with virucide (B). After exposure to virucide, only the most concentrated solution of virus is still able to induce a cytopathic effect. Virus concentrations are denoted as fold dilutions (i.e. 101=10x dilution, 102=100x dilution, etc.)
In an effort to identify novel virucides and citing “a major problem with the testing of virucidal efficacy using current protocols is that scoring of virus-induced cytopathic effect is dependent on subjective visual interpretation using light microscopy,” E.H. Venter and colleagues at the University of Pretoria evaluated the xCELLigence CPE assay as a novel means for testing virucide efficacy.
- A commercially available virucide was incubated at the manufacturer’s recommended concentration with infectious bursal disease virus (IBDV) for 20 minutes.
- The virus was diluted serially and added to confluent Vero cells in E-Plates.
As seen in Figure A, the untreated virus causes a cytopathic effect, with the time of CPE onset being dependent on the virus titer. However, pre-treatment of IBDV with virucide significantly diminished infectivity: whereas the 10x dilution of virus was still able to induce a CPE, 100x and higher dilutions of the virus had no effect on the Vero cells (Figure B).
After evaluating the generated data, the authors concluded, “although the modified [virucide] assay using the xCELLigence system yielded identical results as the traditional [virucide] assay, the system allows virucidal efficacy and cytotoxicity to be measured in a more precise and reproducible fashion.”
Virucide Efficacy Publications
- Ebersohn, K.; Coetzee, P.; Venter, E. H. An Improved Method for Determining Virucidal Efficacy of a Chemical Disinfectant Using an Electrical Impedance Assay. J. Virol. Methods. 2014 Apr, 199, 25–8.
Featured xCELLigence RTCA Systems for Viral CPE Assays
|Dual Purpose||Single Plate||Multi Plate||High Throughput|
|3×16 wells||1×96 wells||6×96 wells||Up to 4×384 wells|