SARS-COV-2 PCR Controls

In the context of SARS-CoV-2 diagnostics, PCR controls are critical components integrated into testing protocols to validate the accuracy, sensitivity, and specificity of the assay. These controls address various stages of the testing process—from sample collection and nucleic acid extraction to amplification and detection—ensuring that results are reliable and free from false positives or negatives.

Types of PCR Controls for SARS-CoV-2 Testing

• Negative Controls:
  • No Template Control (NTC): Contains all PCR reagents except the nucleic acid sample. An NTC checks for contamination in reagents or the environment during setup. Any amplification in this control suggests contamination issues.
• Positive Controls:
  • Synthetic or Inactivated SARS-CoV-2 RNA: A known concentration of SARS-CoV-2 viral RNA or an inactivated virus is included to verify that the assay reagents, primers, and probes can successfully detect the target sequences. This control helps confirm that the PCR setup is functioning correctly.

Application and Importance of Each Control

• Ensuring Specificity and Sensitivity: Positive controls validate that assay conditions (including primer/probe sets and thermal cycling parameters) can accurately detect the SARS-CoV-2 genome at expected sensitivity levels. They help in defining the lower limit of detection (LOD).
• Contamination Monitoring: Negative controls identify any potential contamination events during reagent preparation, sample handling, or assay setup. Detection of amplification in a negative control necessitates investigation and potential repetition of the assay.
• Verification of Nucleic Acid Extraction: Extraction controls like human RNase P or exogenous spiked RNA assure that the sample collection and processing steps yield quality nucleic acids. They help distinguish between true negative results and failures due to poor sample quality or extraction inefficiencies.

By rigorously applying these SARS-CoV-2 PCR controls, laboratories can confidently distinguish true viral presence from assay artifacts, ensuring high diagnostic accuracy essential for patient care and public health surveillance.