Molecular diagnostics in the clinical laboratory

Molecular diagnostics is now established across many hospital and clinical laboratory test menus, supporting applications ranging from infectious disease testing and inherited disease screening to oncology profiling, pharmacogenomics and liquid biopsy. Technologies such as qPCR, digital PCR and NGS enable laboratories to detect clinically relevant molecular targets with increasing precision.

As molecular testing becomes more integrated into routine clinical practice, laboratories are expected not only to generate accurate results but also to demonstrate that analytical claims remain valid throughout the lifetime of the assay. This places greater emphasis on validation, verification, quality control and ongoing performance monitoring.

Quality control for molecular assays

Quality control provides evidence that a molecular assay continues to perform as expected after validation has been completed. Effective control strategies help laboratories identify contamination events, extraction failures, reagent variability and shifts in assay sensitivity before patient results are affected.

Positive controls, negative controls, no-template controls and low-frequency variant controls all contribute to demonstrating that assays remain within established acceptance criteria. Independent quality controls provide an objective assessment of testing performance and may detect issues that are not apparent when relying solely on manufacturer-supplied controls.

Control strategies should reflect assay design and clinical application. Oncology assays, liquid biopsy workflows and minimal residual disease applications often require controls capable of challenging sensitivity claims at clinically relevant variant allele frequencies (VAFs). Infectious disease assays may place greater emphasis on extraction efficiency, inhibition monitoring and contamination detection.

Quality Measurement tools (QMT) can further strengthen quality assurance programmes through trend analysis, peer comparison and centralised review of QC data.

Reference materials for assay validation

Reference materials support assay validation, verification and ongoing quality assurance throughout the molecular testing lifecycle.

During validation studies, characterised reference materials can help establish:

• Analytical sensitivity
• Analytical specificity
• Accuracy
• Precision
• Reproducibility
• Limit of detection
• Reportable range
• Lot-to-lot consistency

For NGS panels, oncology assays and liquid biopsy applications, patient-like and contrived samples can help assess variant detection performance under controlled conditions. As laboratories adopt increasingly sensitive assays, particularly those targeting variants at very low allele frequencies, appropriately characterised reference materials become critical for generating reproducible validation data and supporting analytical claims.

Proficiency testing and external quality assessment

Internal quality control demonstrates day-to-day assay reliability, but it does not show how performance compares with peer laboratories. Proficiency testing and external quality assessment provide independent evaluation of laboratory performance against defined criteria and external benchmarks.

Participation in CAP proficiency testing and EQA programmes can help laboratories identify systematic issues, verify testing consistency and provide objective evidence of competency during accreditation activities. When reviewed alongside internal QC data, these programmes contribute to a comprehensive quality management strategy and support continuous improvement.

Validating laboratory-developed tests

The decision to bring a test in-house is often driven by the need to improve turnaround times, expand testing capabilities or reduce reliance on send-out testing. Before patient testing begins, laboratories must establish and document performance through a structured validation process.

Validation studies typically assess analytical sensitivity, analytical specificity, accuracy, precision, reproducibility and LoD. Depending on assay design and intended use, laboratories may also evaluate reportable range, interference, carryover and workflow robustness.

Reference materials, contrived samples and well-characterised clinical specimens are frequently incorporated into validation studies to provide objective evidence supporting assay claims. Once implementation is complete, ongoing quality control and periodic verification activities help demonstrate continued assay reliability and support inspection readiness.