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Explained: How TrueNat test works

The SARS-CoV-2 virus, which causes Covid-19, does not have a DNA, but an RNA molecule. The reverse transcription process (the RT in RT-PCR) converts the RNA into the DNA molecule before the gene can be captured in the test.

Written by Smita Nair | Panaji |
Updated: June 19, 2020 8:02:34 pm
How TrueNat test works, TrueNat test, Covid test, Covid vaccine, coronavirus vaccine A test sample is loaded onto a TrueNat chip. (Source: Molbio Diagnostics)

A few weeks ago, the Indian Council of Medical Research (ICMR) approved the use of TrueNat machines, manufactured by a Goa-based company, for carrying out confirmatory tests for Covid-19 disease. Before that, these machines, which were originally developed for detecting tuberculosis in patients a few years ago, were being used in the current coronavirus pandemic only for screening patients.

Following the ICMR approval, these machines are now being sought by a number of states, especially those that lack a strong laboratory network that is required to carry out the traditional RT-PCR tests. While Uttar Pradesh has got 117 machines to be deployed in all its 75 districts, Bihar has ordered 50 machines. Jharkhand, Chhattisgarh, Odisha, Telangana, Andhra Pradesh all have sought these machines, and many of these have already been deployed.

How does TrueNat work?

To understand that, it helps to understand how the traditional PCR tests work. PCR is a method to capture a specific gene from the DNA in the swab sample, and multiply it through a series of chemical processes so that it can be detected using fluorescent dyes. Most modern PCR tests, which are used for detecting other kind of viruses as well, work in real time. The result is visible even while the chain reaction is happening.

The SARS-CoV-2 virus, which causes Covid-19, does not have a DNA, but an RNA molecule. The reverse transcription process (the RT in RT-PCR) converts the RNA into the DNA molecule before the gene can be captured in the test.

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TrueNat is a chip-based, battery-operated RT-PCR kit. Initially, it could only identify the E-gene in the SARS-CoV-2 virus. This is the gene that helps the virus build a spherical envelope around it. At this point, TrueNat machines were used as a screening test. Samples that were detected with the E-gene could be sent for confirmatory RT-PCR tests in laboratories. But the new machines are now equipped to detect the RdRp enzyme found in the virus RNA, and hence the ICMR has ruled that these tests can be treated as a confirmation for the presence of the novel coronavirus.

The big difference is that the machine is portable, and tests with it cost much less than the traditional RT-PCR tests. That make it extremely useful for deploying in interior districts and faraway places from where collecting and sending swabs for testing in big cities is a difficult task. In fact, these machines have already been deployed in villages of northern coastal belt of Andhra Pradesh, and inside the forested regions of Gadchiroli district.

How is the test conducted?

Once the swab is collected at kiosks set up at these places, or in containment zones or health camps, it is dipped in a viral transmission medium where it gets neutralized. At the primary healthcare centre or a nearby laboratory, it is transferred to another liquid, a viral lysis bugger, in which the cells break and the impurities are removed. A part of this liquid is then transferred into a cartridge that looks like a flattened tape cassette and is inserted inside a machine very much like a cassette player. The play button activates the process. In just about 20 minutes, the process extracts the RNA, the signature set of genome instructions which command a cell to multiply the virus once it enters a human cell. This RNA extract is then transferred to another machine where the liquid is released into a miniature well that is attached to an electronic chip that is no bigger than a human thumb.

While the miniature well is where the reagent activates the RNA, it is the chip that is fed with all the calculations of the viral load that helps in detecting whether a person is carrying the virus or not. Unlike in the conventional RT-PCR tests, the reagents do not require extreme temperatures in this process, and the quantity of swab required for testing is also much less.

Given that the machines were developed for testing for tuberculosis, did they require adapting for Covid-19 testing?

The machine, designed by Goa-based Molbio Diagnostics Private Limited, was originally developed for the diagnosis of tuberculosis, which affects at least one million children every year. A result of at least a decade of research, these machines had only recently been approved by the World Health Organization (WHO), which attested that these machines “showed similar accuracy” to other WHO approved commercial products for this purpose. These machines were ready to be deployed from April this year.

“That is when Covid happened,” said Shiva Sriram, national sales manager of Molbio Diagnostics, sitting at the Goa manufacturing division. “TB requires early detection, and that is why it is important to have accurate diagnosis. Unlike the smear microscopy which has a 50:50 accuracy, we wanted to attain better accuracy, and kept our research going. It has been 18 years of research…,” he said.

The heart of the machine is designed to detect different kinds of infections. It is the chip that is specific to every infection. The company took 15 days to design the chip which had all the viral load diagnosis for detecting Coronavirus.

According to Sriram, the company was busy making delivery arrangements for the consignee list provided to it by Ministry of Health for TB diagnostics. “Five hundred machines were divided between all the states, and union territories. After that, the states had started placing orders on their own. As of today, about 1,000 machines are already booked, and we are expecting another 3,000 to 4,000 to be ordered in the next two months,” he said.

It was Goa that first purchased the new modified machine for Covid-19 screening. But Andhra Pradesh was the state that used it most extensively, running more than 2.5 lakh screening tests.

The demand grew when the cases began to rise in states like Bihar, Uttar Pradesh and Odisha, as migrants began to return in the second week of May. The machine has already enabled door-to-door surveys in many parts of these states.

How does TrueNat compare with conventional tests?

Conventional RT-PCR requires RNA extraction and analysis to be done in two different rooms backed with cold storage and trained experts handling laboratory designed equipment. TrueNat on the other hand is designed primarily to work at remote locations, and considered as ‘last mile diagnostics’.

“The machine is portable and can be carried in a briefcase, is battery operated with one charge lasting ten hours. An eight-hour shift gives 45 tests and most of the states are currently running three shifts a day,” said Sriram. “Most states have also ordered for the four slot machine which allows four tests to be conducted simultaneously,” he said.

Each of the four slot machines costs upward of Rs 13 lakh but the test kit comes only for about Rs 1,300.

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