Managing a PCR Contamination Event in a Molecular Pathology Laboratory.

Contamination in a molecular laboratory may lead to erroneous results with potential to cause patient harm if not promptly identified and corrected. A general overview of the practices used in molecular laboratories to identify and address contamination once an event has occurred is discussed. The process used to assess the risk associated with the identified contamination event, determine the appropriate course of immediate action, perform a root cause analysis to determine the source of contamination, and assess and document the results of the decontamination process will be reviewed. Finally, the chapter will discuss a return to normal with consideration of appropriate corrective actions to mitigate future contamination events.

SARS-CoV-2 Sequencing for Variant Surveillance.

In this chapter, next-generation sequencing of the entire viral genome of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is described. Successful sequencing of the SARS-CoV-2 virus is dependent upon quality of the specimen, adequate coverage of the entire genome, and up-to-date annotation. Some of the advantages of performing SARS-CoV-2 surveillance using next-generation sequencing are scalability, high-throughput, cost, and full genome analysis. Some of the disadvantages can be expensive instrumentation, large upfront reagent and supply costs, increased time-to-result, computational needs, and complicated bioinformatics. This chapter will provide an overview of a modified FDA Emergency Use Authorization procedure for the genomic sequencing of SARS-CoV-2. The procedure is also referred to as the research use only (RUO) version.

Molecular profiling of vitreous fluid by massively parallel sequencing: a case series.

INTRODUCTION: In cases of suspected intraocular malignancy, vitreous may be the preferred pathologic sample; however, cellularity may be insufficient for definitive cytopathological diagnosis. Ancillary methodology to study vitreous fluid aspiration for mutational analysis may assist in treatment decisions. MATERIALS AND METHODS: Three individual patient vitreous humor samples were received in the laboratory for mutation testing. The samples were collected during standard of care and analyzed for routine cytopathology. In each case, cytopathology was inconclusive and mutational analyses to support diagnostic suspicions were clinically requested. Based on the clinically and pathologically suspected diagnoses, an appropriate massively parallel sequencing assay previously validated for clinical use was performed using DNA extracted from vitreous samples that had previously undergone various processing. Nucleic acid yield was assessed by fluorometric or spectrophotometric methods, with yield ranging from 2.7 to 86.5 ng. Library preparations were performed using standard laboratory protocols. RESULTS: Two of the cases were suspicious for melanoma and a 50-gene solid tumor panel was performed. The third case was worrisome for vitreoretinal lymphoma and a 49-gene myeloid panel was performed. CONCLUSIONS: In all cases, the molecular profiling assisted with the clinical assessment and/or management of each patient.