Epigenetic modifications such as methylation influence cellular differentiation and gene expression. Liquid biopsies are starting to screen for cancer-derived DNA by looking for unexpected epigenetic modifications in circulating cell-free DNA (ccfDNA) that could also be used to assign a tissue of origin to the cancer to direct confirmatory diagnostic procedures. However, because of the low abundance of ccfDNA, obtaining sufficient amounts for assay development, validation, and proficiency testing are difficult. Furthermore, methods of analyzing the epigenetic modifications, such as bisulfite conversion, can damage a significant fraction of the input material; but, failing to carry them out to completion can result in an overestimation of methylation. This makes method optimization very important.

This poster from LGC SeraCare describes the creation and validation of circulating cell-free DNA reference materials with defined amounts of methylation to address the challenges associated with assessing the methylation of circulating cell-free DNA.

Homologous recombination deficiency (HRD) arises due to a defect in DNA repair and serves as an important therapeutic biomarker. NGS assays that measure HRD status via genomic instability are used to stratify ovarian and breast cancer patients and determine eligibility for clinical trials as well as PARP inhibitor and platinum-based therapies. Testing is performed on FFPE biopsied tissue, and, despite the challenges of this sample type and the complexity of analysis, there are currently no commercially available HRD reference materials that enable standardization between assays.

This poster from LGC SeraCare describes the development and validation of reference materials to aid in developing and validating NGS-based HRD assays and for quality control monitoring of genomic instability scores.

Advances in next-generation sequencing are enabling larger panel sizes and increased sensitivities for liquid biopsies. These advances lead to a need for patient-like reference materials with more clinically relevant alterations at lower levels than before. To address this need, LGC SeraCare generated novel circulating tumor DNA (ctDNA) reference materials that combine much of the content of Seraseq ctDNA reference materials and add additional variants and copy number alterations. Because limits of detection of individual mutations are now at or below 0.2 percent variant allele frequency in some ctDNA assays, SeraCare also reduced the lowest VAF from 0.1 percent to 0.01 percent.

This poster from LGC SeraCare provides data on the initial testing of new circulating tumor DNA reference materials that incorporate updated clinically relevant alterations and copy number variants with the lowest variant allele frequency reduced from 0.1 percent to 0.01 percent to accommodate larger NGS panels with increased sensitivities.

Myeloid malignancies are characterized by excessive proliferation, abnormal self-renewal, and differentiation defects of hematopoietic stem cells and myeloid progenitor cells. The detection of these types of malignancies using traditional procedures by obtaining bone marrow samples is invasive and risky. Hence, liquid biopsy-based methods are in high demand to monitor and detect new cases of blood cancer.

This poster from LGC SeraCare summarizes the development of ctDNA myeloid reference materials — a purified mix of 25 myeloid DNA variants that were blended against a wild-type background — for the development, validation, and clinical application of targeted NGS assays as well as digital PCR-based assays that analyze for myeloid diseases in cancer patients.

Single-cell RNA sequencing (scRNA-Seq) has revolutionized basic and translational research by enabling the resolution of distinct cell populations within heterogeneous samples.

This poster from Singular Genomics demonstrates the scRNA-seq capabilities of the G4 Sequencing Platform, which provides users with the flexibility to tailor run sizes and flow cell configurations to the sample set rather than accumulating samples to massively pool onto large flow cells.

Traditional multiplex PCR (mPCR)-based targeted sequencing assays deliver high-sensitivity variant detection with a rapid workflow but are unable to capture gene fusions with novel partners or breakpoints.

This poster from Singular Genomics introduces Ring-Seq, a novel mPCR technology for the targeted detection of gene fusions compatible with traditional mPCR panels, enabling novel fusion detection with minimal panel modification.

As a proof of concept, the authors applied Ring-Seq and the Singular Genomics G4 Sequencing Platform to simultaneously detect clinically relevant translocations and marker VDJ rearrangements of the IGH locus from highly degraded material, thereby resolving a key weakness of traditional B-cell clonality assays.

Next-generation sequencing has become an indispensable tool for the diagnosis of genetic disease, though there remains a need to reduce turnaround for time-sensitive applications. Reducing turnaround requires faster sequencing and accelerated data analysis. We recently introduced the Singular Genomics G4 Sequencing Platform for rapid sequencing by synthesis, which can deliver four human whole genomes at ~30x coverage in under 19 hours.

This poster from Singular Genomics discusses the need to reduce sequencing turnaround for time-sensitive applications and presents accelerated pipelines for whole-exome germline and targeted somatic variant detection on the G4 Sequencing Platform that leverage the Nvidia Clara Parabricks platform.

Next-generation sequencing has become a foundational tool for both biological research and in vitro diagnostics, particularly in oncology, immunology, and the detection of genetic disorders. Despite its success, there is a need for new DNA sequencing platforms that combine high accuracy, speed, and flexible throughput to provide timely results and cost-effective operations for research and clinical applications.

This poster from Singular Genomics presents a performance evaluation of the novel Singular Genomics G4 Sequencing Platform for rapid sequencing by synthesis, demonstrating its utility for whole-genome sequencing, bulk and single-cell RNA-seq, and high-fidelity demultiplexing via unique dual indices.

Next-generation sequencing (NGS) has enabled rapid progress in cancer diagnostics. However, challenges remain around the detection of somatic variants at allele frequencies similar to or below the error rate of on-market DNA sequencers (typically a ~10^-3) when using standard library preparation methodologies.

This poster from Singular Genomics introduces HD-Seq, a novel library preparation and sequencing method that relies on linking the original paired strands in a dsDNA fragment and that allows for efficient and high-accuracy sequencing, achieving error rates in the range of ~10^-5-10^-6.

In neuroscience research, it is critical to study the complex structure of the different cell types in the brain, as the chemical, morphological, and functional features of brain cell types are extremely diverse.

NanoString’s nCounter Analysis System can look at the expression of up to 800 genes associated with neuropathology and neuroinflammation. These panels examine and identify five different CNS cell types and 14 different immune cell types to gather a signature profile of biological processes and diseases.

This application note from Canopy Biosciences, a Center of Excellence for NanoString services, describes two recent neuroscience studies using the nCounter platform: one that classifies pediatric ependymomas using custom antibody probes, and another that investigates miRNA expression in microglial-derived extracellular vesicles in Alzheimer’s disease.

Predictive biomarkers help to identify tumors that are most likely to benefit from a specific treatment and spare patients from toxic effects of ineffective therapies. Both genetic mutations and protein levels in diseased tissues are examples of predictive biomarkers, yet few platforms enable the simultaneous evaluation of both RNA and protein.

The GeoMx Digital Spatial Profiler (DSP) from NanoString provides morphological context in spatial transcriptomics and spatial proteomics experiments for immuno-oncology research. The GeoMx DSP platform can investigate novel biomarkers in tissue samples because it offers digital profiling of protein and RNA analytes on the same slide.

This application note from Canopy Biosciences, a Center of Excellence for NanoString services, outlines a recent study that used the GeoMx DSP to investigate novel biomarkers associated with beneficial PD-1 therapy in formalin-fixed, paraffin-embedded samples from patients with non-small cell lung cancer.

Next-generation sequencing (NGS) oncology panels are becoming integral in hospital and academic settings to guide patient treatment and enrollment in clinical trials. But as NGS is increasingly used in precision oncology and an unprecedented amount of genomic information is uncovered, there is an industry-wide issue of standardization: a high degree of variability in variant interpretation currently exists across laboratories.

Inconsistencies in variant interpretation among laboratories can create confusion for clinicians and their patients, leaving them unable to confidently use genetic information to manage healthcare decisions. In this live panel discussion, experts in NGS testing and clinical informatics explore the issues surrounding the standardization of variant interpretation and consider how interpretation guidelines and clinical decision support (CDS) software can help to mitigate variability between laboratories.

Topics of discussion include:

• Findings of a peer-reviewed study comparing the accuracy and consistency of variant assessments from commercial CDS software to the internal variant interpretation methods of eight laboratories.
• Causes of inter-laboratory inconsistency in variant interpretation and how to mitigate variability.
• The benefits of having an up-to-date, high-quality knowledge base to produce consistent, evidence-based clinical classifications.

This video from Asuragen presents data from a method comparison between the QuantideX qPCR BCR-ABL IS kit for chronic myeloid leukemia monitoring and another commercially available CE-marked BCR-ABL1 quantification kit for monitoring BCR-ABL major breakpoints.

Sex chromosome aneuploidies (SCAs) are genetic conditions caused by abnormalities in the number of X and Y sex chromosomes, which include 45, X (Turner syndrome), 47, XXY (Klinefelter syndrome), and 47, XYY (Jacobs syndrome). With a combined incidence of ~1 in 400 live births, SCAs occur more frequently than trisomy 21 (1:700). The majority of commercially available cell-free DNA (cfDNA) screens, also known as non-invasive prenatal tests (NIPT), can provide optional sex aneuploidy analysis.

Guidelines recommend the use of external run controls to validate test performance for SCAs when NIPTs are first implemented in a clinical lab and for routine performance monitoring. However, this can be problematic due to the rarity, low volume, and ethical considerations in obtaining clinical samples. This is especially important as current NIPT technologies have a lower sensitivity and positive predictive value for SCAs as compared to trisomy 21.

This poster from LGC SeraCare presents the development, characterization, and clinical performance of novel SNP-matched sex chromosome aneuploidy reference materials on several commercial NIPT assays.