Case study: Demonstrating the mechanism of action of a rhinovirus inhibitor using a virus-binding assay
Scientific challenge
Quantifying neutralising activity
A client developing antiviral and antibody-based interventions against measles virus required a robust method for quantifying neutralising activity.
VRS had previously developed and established a microneutralisation assay using an immunofluorescence-based readout, in which infection was quantified by measuring the number of infected cells. This approach provided a practical and scalable alternative to traditional plaque-based methods and was used throughout the study to characterise neutralising activity.
At the end of the programme, the client wanted to establish how closely the VRS assay compared with the plaque reduction neutralisation test (PRNT), widely regarded as the gold-standard method for measuring measles virus neutralisation.
Virus |
Measles virus |
Project |
Comparison of a high-throughput microneutralisation assay with the plaque reduction neutralisation test (PRNT) |
Assay |
Immunofluorescence-based microneutralisation assay, PRNT/FRNT |
Client |
Biopharmaceutical company |
Stage |
Antiviral and antibody development |
Services |
Bespoke assay development, microneutralisation assay, immunofluorescence, PRNT comparison, assay validation |
Related expertise |
Measles virus, neutralisation assays, immunofluorescence, foci-based assays, assay comparison |
Key considerations
Adapting an established foci-based assay
The comparison needed to demonstrate that the two assays generated comparable measurements of neutralising activity, rather than simply showing that both assays could distinguish positive and negative samples.
Key considerations included:
- adapting the established VRS foci-based assay specifically for measles virus,
- comparing the two methods using a representative panel of positive and negative quality control samples,
- performing the assays side by side under controlled conditions,
- assessing agreement between the resulting neutralisation profiles and IC50 values.
A close correspondence between the two methods would provide confidence that the higher-throughput immunofluorescence assay was measuring the same underlying biological activity as the established PRNT.
Our Approach
Directly compare two approaches
VRS first adapted its established 96-well foci-based PRNT assay to measles virus.
To directly compare the two approaches, VRS designed a one-off comparative study in which a panel of positive and negative QC samples was tested side by side using both the immunofluorescence-based microneutralisation assay and PRNT.
The same samples were therefore assessed using two fundamentally different readout approaches: infected-cell quantification by immunofluorescence and plaque reduction by the conventional PRNT method.
Neutralisation curves were generated for the samples tested, and the resulting IC50 values were compared between the two assays.
Outcome
Practical advantages of a 96-well format
The two assays showed complete correspondence across the QC sample panel, with positive and negative samples demonstrating the expected profiles in both methods.
Importantly, the IC50 values generated using the VRS immunofluorescence-based microneutralisation assay closely matched those obtained using PRNT, demonstrating strong agreement between the two approaches.
The comparison provided the client with confidence that the established VRS assay was accurately measuring measles virus neutralising activity while offering the practical advantages of a 96-well, immunofluorescence-based format.
VRS INSIGHT
Finding a robust and scalable alternative
Different neutralisation assays can use very different experimental readouts while measuring the same underlying biological phenomenon. Demonstrating equivalence between methods can therefore be particularly valuable when a project uses a higher-throughput assay but requires confidence against an established reference method.
In this case, direct side-by-side comparison with PRNT demonstrated that the VRS immunofluorescence-based microneutralisation assay generated highly comparable neutralisation measurements, supporting its use as a robust and scalable alternative for measles virus studies.
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