The Centre for Cellular and Molecular Biology (CCMB), a Hyderabad-based laboratory under the Council of Scientific & Industrial Research (CSIR), is in the news for the development of a new test for SARS-CoV-2, the virus that causes COVID-19. This test, named ‘Reverse Transcription nested Polymerase Chain Reaction’ (RT-nPCR), is significantly different from the recommended RT-qPCR testing method, with some key advantages that make it more accessible and potentially more reliable. This article explores the details of this development and its implications for COVID-19 testing.
Introduction to the RT-nPCR Test
Unlike the currently approved RT-qPCR testing method, the RT-nPCR test does not rely on real-time monitoring of DNA amplification; instead, it utilises standard RT-PCR as part of an endpoint assay to measure the biochemical activity of collected samples. This innovation was designed keeping in mind cost-effectiveness and ease of use. The test makes use of a low-tech model, making it affordable and user-friendly. However, this novel method is still pending approval from the Indian Council of Medical Research (ICMR).
Understanding the ICMR Recommended RT-qPCR Test
The ICMR recommends the Reverse Transcription Polymerase Chain Reaction (RT-qPCR) test for detecting the novel coronavirus. This test uses an enzymatic reaction to amplify deoxyribonucleic acid (DNA). Unlike traditional PCR methods that depend on end point analysis, RT-qPCR allows real-time monitoring of DNA amplification, enabling quantification of DNA.
However, since the coronavirus structure consists of Ribonucleic Acid (RNA) rather than DNA, the RNA is converted into DNA using a technique called Reverse Transcription (RT). This is a crucial step for detection of coronavirus through this method.
Comparing RT-qPCR and RT-nPCR
The most significant difference between RT-qPCR and the newly developed RT-nPCR lies in what is being studied. In RT-qPCR, the viral RNA is quantified; conversely, RT-nPCR studies the viral RNA that nests.
Furthermore, RT-qPCR testing has several challenges: it’s costly, time-consuming, and requires sophisticated equipment including a pricey real-time thermal cycler. On the other hand, RT-nPCR is economical, enables large-scale testing, and doesn’t necessitate any special apparatus. It only requires a simple PCR machine and can be conducted in a basic lab set-up.
Effectiveness of the RT-nPCR Test
When comparing the two methods, the CCMB scientists noticed a high percentage of false negatives with RT-qPCR testing. In contrast, the RT-nPCR test identified 90% of detected samples as positive — aligning with RT-qPCR outcomes — proving its reliability. Additionally, RT-nPCR identified 13% of samples as positive among those that were negative according to the standard RT-qPCR test, indicating a potential detection of likely false negatives.
Overall, the RT-nPCR test demonstrates promise as a cost-effective, efficient and reliable addition to current COVID-19 testing methods, pending approval from ICMR.
