Compliant Application and Performance Validation of NGS Library Preparation Kits in Clinical Testing

I. Classification and Compliance Prerequisites for NGS Library Preparation Kits in Clinical Settings

In the field of clinical molecular diagnostics, the use of NGS library preparation kits should be managed in accordance with the intended use of the product and its regulatory classification. If a kit is intended as a component or accessory of an in vitro diagnostic reagent, attention should be paid to its filing or registration status; if a laboratory adopts a laboratory-developed method for testing, a rigorous performance verification and quality management system must be established. In accordance with the general requirements of China's medical device regulations, kit instructions, labels, and lot information should be clear and traceable, and manufacturers should possess corresponding production and quality assurance capabilities. Before introduction, clinical laboratories should verify product compliance documents to avoid using research-grade reagents directly for clinical reporting. In addition, attention should be paid to the intended use, sample types, and compatible instrument ranges stated in the kit instructions to ensure consistency with the laboratory's actual testing items.

II. Core Indicators and General Methods in Performance Verification

NGS library preparation kits typically require performance verification before clinical testing, with common indicators including precision, accuracy, analytical sensitivity, specificity, and reproducibility. The verification protocol may refer to relevant technical guidelines, selecting representative sample types and mutation frequency ranges for intra-run and inter-run testing. In the library preparation step, particular attention should be paid to the detection stability of low-abundance variants and the coverage uniformity of high-GC regions and repetitive sequence regions. During verification, manufacturer-promoted parameters should not be simply adopted; instead, independent confirmation should be performed under the laboratory's own conditions, with written verification records generated. For different nucleic acid input amounts, different degrees of fragmentation, and different library preparation lots, verification experiments should be designed separately to fully evaluate the performance of NGS library preparation kits in real samples.

III. Key Points of Sample Processing and Whole-Process Quality Control

The nucleic acid quality of clinical samples directly affects the final performance of NGS library preparation kits. Common samples include formalin-fixed paraffin-embedded tissue, peripheral blood, and fresh tissue, which differ in extraction, fragmentation, repair, and amplification steps. Laboratories should establish sample acceptance criteria, nucleic acid quantification and integrity assessment procedures, and set quality control nodes before and after library preparation. For example, for samples with library concentrations below the routine threshold, it should be assessed whether to proceed with sequencing or re-perform library preparation; for libraries with adapter contamination or abnormal fragment distribution, the causes should be analyzed and corrective measures taken. Whole-process quality control helps reduce the risk of testing failure and result misinterpretation. At the same time, retention sample spot checks should be periodically performed on library preparation kits to monitor lot-to-lot quality fluctuations.

IV. Boundaries of Compliant Use and Report Interpretation

NGS library preparation kits are only one part of the entire testing workflow, and their stable quality cannot be equated with reliable clinical conclusions. When issuing reports, laboratories should make judgments based on comprehensive consideration of sample information, sequencing data quality, bioinformatics analysis workflows, and clinical significance. For tumor-related gene variants, results should be positioned as reference information to assist clinical decision-making, and specific diagnosis and treatment plans should be comprehensively determined by clinicians in conjunction with the patient's condition. Meanwhile, relevant institutions should continuously carry out internal quality control and external quality assessment, maintain personnel training and document updates, and ensure that NGS library preparation kits operate stably within a compliant framework. Any improvement in kit performance or workflow adjustment should be implemented only after evaluation through a change control procedure.