Archer exhibits at USCAP 2017 Annual Meeting

R. Kulesus | 17 Jan, 2017

March 4 - 10 | Booth #238

San Antonio, TX

Stop by booth 238 and pick up a 2017 poster calendar!

Get the poster!

Learn more about how Archer® Anchored Multiplex PCR (AMP™) enrichment chemistry harnesses the power of next-generation sequencing (NGS).

The United States and Canadian Academy of Pathology (USCAP) annual meeting boasts the largest and most influential world meeting of pathologists.

USCAP has been the most prestigious provider of continuing medical education (CME) for pathologists globally for more than a century

Poster abstracts

Sensitive and Specific Detection of Mutations in Circulating Tumor DNA with Anchored Multiplex PCR and Next-Generation Sequencing {9915}

Sensitive and Specific Detection of Mutations in Circulating Tumor DNA with Anchored Multiplex PCR and Next-Generation Sequencing {9915}

Jerome E. Lee, Josh D. Haimes, Namitha Manoj, Eric M. Davis, Ian McKittrick, Katelyn E. Trifilo, Laura A. Johnson, Laura M. Griffin, Brian A. Kudlow, and Brady P. Culver

ArcherDX, Boulder, CO USA

Background: Liquid biopsies are a promising, minimally invasive alternative to solid tumor biopsies that have potential cost, time and safety benefits, as well as a greater ability to interrogate heterogeneous tumors. However, except in advanced disease states, cell free DNA (cfDNA) is typically of low abundance and only a small portion of cfDNA originates from tumor cells as circulating tumor DNA (ctDNA), which tends to be highly fragmented (100-300bp). Therefore, NGS-based assays to detect variants in ctDNA must be sensitive enough to detect mutations at allele frequencies (AF) 2% from 100ng of highly fragmented DNA.

Design: We developed the Archer® Reveal ctDNA™ 28 assay based on Anchored Multiplex PCR (AMP™), a target enrichment method for NGS that uses unidirectional gene-specific primers and molecular barcoded (MBC) adapters for amplification. This enables preferential enrichment of small, degraded ctDNA fragments over long genomic DNA fragments, thereby increasing detection sensitivity of mutations in ctDNA by NGS. MBC adapters ligated prior to amplification permit post-sequencing error correction, reducing background noise and increasing analytical sensitivity of detecting ultra low-allele frequency variants. Variant filtering in the Archer Analysis pipeline further increases the specificity of variant calls.

Results: Using commercially available synthetic ctDNA standards, we demonstrate that the AMP-based Reveal ctDNA 28 assay preferentially enriches small, fragmented ctDNA over genomic DNA. Based on sequenced reads, AMP enabled interrogation of more than 65% of the input molecules from 50ng starting material. As a result, we show 100% detection sensitivity for 1% AF variants using 10ng DNA input and 71.9% detection sensitivity for 0.1% AF variants using 50ng DNA input. MBC-enabled post-sequencing error correction and variant filtering reduced the number of false positives by 98%, resulting in 91.7% specificity. Finally, Reveal ctDNA 28 accurately detected mutations present in corresponding tissue biopsies with under 10ng liquid biopsy-derived DNA.

Conclusions: The Archer Reveal ctDNA 28 assay is a powerful tool for sensitive and specific NGS-based detection of variants in reference ctDNA standards and shows promise for accurate detection of variants in ctDNA from clinical liquid biopsies.

B- and T-cell immune repertoire characterization by Anchored Multiplex PCR and next-generation sequencing {13776}

B- and T-cell immune repertoire characterization by Anchored Multiplex PCR and next-generation sequencing {13776}

Jens Eberlein, Thomas Harrison, Ian McKittrick, Megan Wemmer, Laura M. Griffin, Brady P. Culver, Laura Johnson, Brian A. Kudlow

ArcherDX, Boulder, CO USA

Background: The adaptive immune system is involved in various disease conditions including cancer, chronic infection, autoimmune disease and transplant rejection. Adaptive immunity is mediated by B and T lymphocytes, which are activated upon antigen binding to antigen receptors expressed on their surface. Therefore, the spectrum of these antigen receptors, or immune repertoire (IR), provides a means to monitor adaptive immune responses to disease, vaccination and therapeutic interventions. Next-generation sequencing (NGS) of antigen receptor genes is a valuable tool in the study of disease states and responses to various interventions. Traditional amplicon-based NGS assays use opposing primers for targeted amplification of rearranged antigen receptor genes. Thus, large primer panels are required to capture the extensive combinatorial diversity exhibited by the IR. Quantification from such assays requires a complex system of synthetic controls to account for differential amplification efficiency across segment combinations. Here, we describe an Anchored Multiplex PCR (AMP)-based NGS assay to analyze the IR, employing a minimal set of gene-specific primers in conjunction with molecular barcodes (MBCs) to reduce amplification bias.

Design: AMP uses MBCs ligated to cDNA ends and gene-specific primers for amplification, enabling immune chain mRNA interrogation from a single side. This eliminates the need for opposing primers that bind within the highly variable V-segment, eliminating clone dropout due to somatic hypermutation. Furthermore, this facilitates CDR3 sequence capture from highly fragmented RNA inputs.

Results: We validated the quantitative reproducibility and sensitivity of the AMP-based IGH assay using mRNA isolated from peripheral blood leukocytes of healthy and B-cell chronic lymphocytic leukemia (B-CLL) donors. Our data showed high reproducibility between replicates and quantitative clone tracking down to 0.01%, with the ability to determine IGHV mutational status. We also validated the quantitative reproducibility and sensitivity of the AMP-based T-cell receptor (TCR) assay using high-quality mRNA isolated from peripheral blood leukocytes and highly fragmented RNA isolated from formalin-fixed paraffin-embedded (FFPE) samples. Our data indicate that clonal diversity in sequencing data is driven by input quantity, total T-cell number, and, to a lesser degree, mRNA quality.

Conclusions: AMP-based NGS with MBC quantification and error-correction is a powerful method to characterize the immune repertoire.

Archer's newest assays

Reveal ctDNA Kit

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Mutation profiling from circulating tumor DNA in liquid biopsies

The Archer® Reveal ctDNA™ 28 Kit for Illumina® is an advanced and user-friendly solution for targeted NGS of circulating cell-free tumor DNA e.g., ctDNA, ccfDNA, cfDNA from 28 genes commonly found mutated in solid tumor type cancers.

Immunoverse Kit

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Immune repertoire sequencing assays

Archer® Immunoverse™ kits are targeted NGS assays to characterize the human immune repertoire from RNA input. Powered by AMP, the lyophilized kits uniquely tag and amplify V(D)J rearrangements for sequencing on Illumina® platforms.


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Comprehensive mutation profiling for cystic fibrosis

The Archer® VariantPlex® CFTR kit is a targeted NGS assay for comprehensive detection of known and unknown variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene.

About ArcherDX

ArcherDX addresses the bottlenecks associated with using NGS by offering a robust platform for targeted sequencing applications.

Archer target enrichment assays utilize AMP chemistry to generate highly enriched sequencing libraries for comprehensive profiling of fusions, CNVs, SNVs and indels. Amplification from independent, unidirectional primers and universal molecular barcoded adapters permit identification of novel gene fusions and mutations with nucleotide-level resolution. Requiring only one intact primer-binding site, AMP chemistry is uniquely suited to amplify small, degraded fragments, enabling solid tumor mutation profiling from FFPE samples and liquid biopsies. Archer’s easy-to-use, lyophilized kits generate sequencing-ready libraries from RNA, DNA, and liquid biopsy-derived ctDNA. Complemented by the Archer suite of assay design and bioinformatics analysis, Archer’s FusionPlex, VariantPlex and Reveal ctDNA assays facilitate complex mutation identification and discovery, while Immunoverse assays enable quantitative profiling of the expressed immune repertoire.