The Real-time PCR marketplace continues to rapidly increase. The SARS-CoV-2 pandemic contributed to the drive for quicker, high-yield tests.
Improved RT-PCR methods employ low reagent intake along with greater reproducibility and precision. These machines automate the workflow and use real-time monitoring. They also offer precision and customization.
Polymerase chain reaction (PCR) represents a pivotal step forward in biotechnology. It’s now a vital tool in test kits and solutions within the healthcare industry. Keep reading to learn more about RT-PCR’s role in infectious disease testing.
PCR vs qPCR in Infectious Disease Testing
The PCR process including qPCR/RT-PCR biotechnologically amplifies DNA for different purposes. PCR is beneficial for bacteria, viruses, and other pathogens containing DNA for amplification. If you must transcribe RNA to DNA for amplification, RT-PCR/qPCR processes are best.
PCR testing involves a qualitative technique versus the quantitative method used for RT-PCR. PCR returns a result describing if the target DNA sequence is present or absent. PCR works well for diagnosing African swine fever, Ebola, and foot-and-mouth disease.
During RT-PCR testing, the machine heats and cools the mixture. This induces precise chemical reactions to generate new, identical copies of viral DNA’s target segments. These copies double with each cycle.
Standard RT-PCR’s first step of phase one subjects the samples to 50.0°C for two minutes. This establishes temperature equilibrium and prepares the samples to react.
The second step serves to activate the polymerase in the PCR mix. The temperature is increased to 95.0°C for ten minutes.
Next, the sample undergoes phase two, which stimulates amplification:
- Denaturing phase: 15 seconds at 95.0°C
- Annealing and elongation phase: 1 minute at 60.0°C
The machine repeats this second phase for 40 cycles. Marker labels attach to each new DNA copy strand as it’s built.
The marker releases a fluorescent dye that the machine’s computer tracks. Real-time data displays on the monitor. Once fluorescence levels pass a set threshold, the virus’ presence is confirmed.
Application Of Real-Time PCR
The RT-PCR test is clinically used to diagnose viral diseases. With the COVID-19 pandemic, the need for rapid detection became vital. Faster diagnosis helped manage limited isolation beds and critically ill patients at the beginning of the pandemic.
RT-PCR coronavirus testing detects SARS-CoV-2 viral nucleic acids or proteins. This test offers high specificity for the presence of this viral nucleic acid.
Yet it can’t differentiate between a viable and non-viable virus. Thus, the test can’t determine if the person is currently infectious. This impacted isolation requirements.
RT-PCR Thermal Cyclers
Two brands of RT-PCR Thermal Cyclers include QIAquant instruments and qTOWER³. They offer peerless temperature control precision thermal blocks regardless of the number of samples. You get a fast, sensitive detection using high-performance optics.
You’re guaranteed homogenous illumination and excitation of each sample. These machines use proven filter modules to achieve unique flexibility with each cycle. You can also expand and configure these modules to enable up to six-fold multiplexing.
QIAquant instruments reduce processing times by completing 40 cycles in under 30 minutes. This is accomplished using rapid heating and cooling ramp rates. It also employs high-quality fiber-optic shuttles.
The qTOWER³ 84 has a 16-fiber high-performance optic as well. You’ll get results for a full 348 well plate in six seconds regardless of the number of filters used.
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