OVERVIEW DATA ORDERING INFORMATION RESOURCES
OVERVIEW

High Sensitivity Synthetic Reference Material for Liquid Biopsy Assay Development and Process Control

The Twist cfDNA Pan-Cancer Reference Standard v2 is a comprehensive tool created to assist researchers in both the development and quality monitoring of clinically relevant variants for developing NGS-based assays. Additionally, researchers can utilize these reference standards to define two essential analytical parameters: the Limit of Detection (LoD) and the Limit of Blank (LoB) for their assays. Composed of wild-type (WT) background cell-free DNA (cfDNA) from a cell-line and synthetic oligos carrying mutant alleles, this reference material offers the precision and sensitivity needed for researching liquid biopsy assay development. 

Broad Variant Coverage

The Twist cfDNA Pan-Cancer Reference Standard v2 offers an extensive array of circulating tumor DNA (ctDNA) variants. These variants closely mirror the size and distribution of cfDNA fragments, which can make it an indispensable tool for accurately simulating real-world scenarios. It comprises synthetically-printed variants, including single nucleotide variants (SNVs), insertion-deletions (INDELs), and structural variants (SVs).

Comprehensive Variant Sites

With over 400 variant sites spanning 84 genes, including literature-curated, clinically-relevant variants, this reference standard promotes a comprehensive approach to cfDNA analysis. It empowers researchers to detect a wide range of genetic variations associated with cancer.

Empower LoD and LoB Establishment

The Twist cfDNA Pan-Cancer Reference Standard v2 is meticulously designed to assist in determining the analytical Limit of Detection (LoD) and the Limit of Blank (LoB) of your assays. This critical capability can enable you to define the lowest detectable levels of target variants and establish a baseline for assay sensitivity.

Low Background Error Rate

Comparative analyses demonstrate that the Twist cfDNA Pan-Cancer Reference Standard v2 exhibits a significantly lower error rate in comparison to its predecessor (v1) and other competing products. 

Compatibility and Versatility

The Twist cfDNA Pan-Cancer Reference Standard v2 standard seamlessly integrates with Twist’s suite of target enrichment systems and custom capture panels.  The Twist cfDNA Pan-Cancer Reference Standard v2 Set consists of 7 individual tubes with 300 ng per tube for each of the 7 different VAFs: 0% (WT), 0.1%, 0.25%, 0.5%, 1%, 2%, 5%. Twist also offers each individual VAF in a smaller 600ng and larger 3ug configurations.

Comprehensive Detection of Cancer Variants
Comprehensive Detection of Cancer Variants
456 unique naturally occurring cancer variants
132 clinically relevant variants
Covers 84 different genes involved in cancer
Exceptional Sensitivity of Background DNA
Exceptional Sensitivity of Background DNA
Background cfDNA derived from non-amplified cell-line DNA
DNA size profile and post-sequencing profile mimic native cfDNA
Designed for Precision and Flexibility
Designed for Precision and Flexibility
Seven individual variant allele percentages to choose from in large and small configurations
Digital Droplet PCR verification of VAF percentages
Convenient test set of all VAF percentages available

*For research use only. Not for use in any diagnostic or clinical procedures.

High Sensitivity Synthetic Reference Material for Liquid Biopsy Assay Development and Process Control

The Twist cfDNA Pan-Cancer Reference Standard v2 is a comprehensive tool created to assist researchers in both the development and quality monitoring of clinically relevant variants for developing NGS-based assays. Additionally, researchers can utilize these reference standards to define two essential analytical parameters: the Limit of Detection (LoD) and the Limit of Blank (LoB) for their assays. Composed of wild-type (WT) background cell-free DNA (cfDNA) from a cell-line and synthetic oligos carrying mutant alleles, this reference material offers the precision and sensitivity needed for researching liquid biopsy assay development. 

Broad Variant Coverage

The Twist cfDNA Pan-Cancer Reference Standard v2 offers an extensive array of circulating tumor DNA (ctDNA) variants. These variants closely mirror the size and distribution of cfDNA fragments, which can make it an indispensable tool for accurately simulating real-world scenarios. It comprises synthetically-printed variants, including single nucleotide variants (SNVs), insertion-deletions (INDELs), and structural variants (SVs).

Comprehensive Variant Sites

With over 400 variant sites spanning 84 genes, including literature-curated, clinically-relevant variants, this reference standard promotes a comprehensive approach to cfDNA analysis. It empowers researchers to detect a wide range of genetic variations associated with cancer.

Empower LoD and LoB Establishment

The Twist cfDNA Pan-Cancer Reference Standard v2 is meticulously designed to assist in determining the analytical Limit of Detection (LoD) and the Limit of Blank (LoB) of your assays. This critical capability can enable you to define the lowest detectable levels of target variants and establish a baseline for assay sensitivity.

Low Background Error Rate

Comparative analyses demonstrate that the Twist cfDNA Pan-Cancer Reference Standard v2 exhibits a significantly lower error rate in comparison to its predecessor (v1) and other competing products. 

Compatibility and Versatility

The Twist cfDNA Pan-Cancer Reference Standard v2 standard seamlessly integrates with Twist’s suite of target enrichment systems and custom capture panels.  The Twist cfDNA Pan-Cancer Reference Standard v2 Set consists of 7 individual tubes with 300 ng per tube for each of the 7 different VAFs: 0% (WT), 0.1%, 0.25%, 0.5%, 1%, 2%, 5%. Twist also offers each individual VAF in a smaller 600ng and larger 3ug configurations.

Comprehensive Detection of Cancer Variants
Comprehensive Detection of Cancer Variants
456 unique naturally occurring cancer variants
132 clinically relevant variants
Covers 84 different genes involved in cancer
Exceptional Sensitivity of Background DNA
Exceptional Sensitivity of Background DNA
Background cfDNA derived from non-amplified cell-line DNA
DNA size profile and post-sequencing profile mimic native cfDNA
Designed for Precision and Flexibility
Designed for Precision and Flexibility
Seven individual variant allele percentages to choose from in large and small configurations
Digital Droplet PCR verification of VAF percentages
Convenient test set of all VAF percentages available

*For research use only. Not for use in any diagnostic or clinical procedures.

DATA
Fragment Profile Comparison of Twist’s Reference Standards and Competitors

DNA fragment analysis of the Twist cfDNA Pan-cancer Reference Standard v2, native cfDNA and competitor reference standards equalized for peak maximum height. Twist standards have a size distribution that emulates native cfDNA including a prominent mononucleosomal peak and dinucleosomal peak.

comparison
Schematic of synthetic variant DNA design

Variant DNA molecules tile across the genomic locus of the variant (purple dashed line with star) with extensive overlap, providing high coverage, fragment diversity, and location diversity of the variant site relative to the termini of the DNA molecule. 

 

Schematic
Background Mutation Rate Comparison

Analysis of background mutation rates is defined as the rate of non-germline variants detected in duplex consensus reads from a target-enriched WT sample, and was performed across Twist cfDNA v2 standard, native cfDNA, a competing DNA standard, and the Twist cfDNA v1 standard. The Twist Pan-Cancer v2 background error rate is similar to native cfDNA and lower than the previous v1 and competitor product due to a high-fidelity production process.

 

Background Mutation Rate Comparison
Indel size distribution

Visualization of the insertion or deletion size of the included INDELs in the reference standards via a histogram of binwidth 1, where negative numbers are deletions and positive numbers are insertions. SNVs are not shown. Included INDELs sample a wide range of sizes, allowing for broad analysis of INDEL detection performance. INDELs range from deletions of 30 bp and insertions of 15 bp.I was

Indel size distribution
Variant Calling in Target Enrichment Workflow

Average variant allele frequency (VAF) detected by NGS in different reference standard VAFs (shown on a log scale). The Twist cfDNA Pan-Cancer Reference Standards were captured with a custom panel targeting 215 single nucleotide variants (SNVs) found in the standards and sequenced to 80,000X raw coverage before UMI deduplication. Detected mean VAFs from NGS mirror closely the intended VAF across all levels tested, facilitating easy assay benchmarking.

 

Variant Calling
Fragment Profile Comparison of Twist’s Reference Standards and Competitors

DNA fragment analysis of the Twist cfDNA Pan-cancer Reference Standard v2, native cfDNA and competitor reference standards equalized for peak maximum height. Twist standards have a size distribution that emulates native cfDNA including a prominent mononucleosomal peak and dinucleosomal peak.

comparison
Schematic of synthetic variant DNA design

Variant DNA molecules tile across the genomic locus of the variant (purple dashed line with star) with extensive overlap, providing high coverage, fragment diversity, and location diversity of the variant site relative to the termini of the DNA molecule. 

 

Schematic
Background Mutation Rate Comparison

Analysis of background mutation rates is defined as the rate of non-germline variants detected in duplex consensus reads from a target-enriched WT sample, and was performed across Twist cfDNA v2 standard, native cfDNA, a competing DNA standard, and the Twist cfDNA v1 standard. The Twist Pan-Cancer v2 background error rate is similar to native cfDNA and lower than the previous v1 and competitor product due to a high-fidelity production process.

 

Background Mutation Rate Comparison
Indel size distribution

Visualization of the insertion or deletion size of the included INDELs in the reference standards via a histogram of binwidth 1, where negative numbers are deletions and positive numbers are insertions. SNVs are not shown. Included INDELs sample a wide range of sizes, allowing for broad analysis of INDEL detection performance. INDELs range from deletions of 30 bp and insertions of 15 bp.I was

Indel size distribution
Variant Calling in Target Enrichment Workflow

Average variant allele frequency (VAF) detected by NGS in different reference standard VAFs (shown on a log scale). The Twist cfDNA Pan-Cancer Reference Standards were captured with a custom panel targeting 215 single nucleotide variants (SNVs) found in the standards and sequenced to 80,000X raw coverage before UMI deduplication. Detected mean VAFs from NGS mirror closely the intended VAF across all levels tested, facilitating easy assay benchmarking.

 

Variant Calling
ORDERING INFORMATION
PRODUCT SKUS

107576

Twist cfDNA Pan-Cancer Reference Standard v2 Set, 300ng kit

107577

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0% (WT), 3 ug

107578

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.1%, 3 ug

107579

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.25%, 3 ug

107580

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.5%, 3 ug

107581

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 1%, 3 ug

107582

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 2%, 3 ug

107583

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 5%, 3 ug

107584

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0%, 600ng

107585

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.1%, 600ng

107586

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.25%,600ng

107587

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.5%, 600ng

107588

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 1%, 600ng

107589

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 2%, 600ng

107590

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 5%, 600ng

*Kit includes 7 VAFs individually: 0% (WT), 0.1%, 0.25%, 0.5%, 1%, 2%, 5% at 300ng each tube

*For research use only. Not for use in any diagnostic or clinical procedures.

PRODUCT SKUS

107576

Twist cfDNA Pan-Cancer Reference Standard v2 Set, 300ng kit

107577

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0% (WT), 3 ug

107578

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.1%, 3 ug

107579

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.25%, 3 ug

107580

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.5%, 3 ug

107581

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 1%, 3 ug

107582

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 2%, 3 ug

107583

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 5%, 3 ug

107584

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0%, 600ng

107585

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.1%, 600ng

107586

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.25%,600ng

107587

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 0.5%, 600ng

107588

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 1%, 600ng

107589

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 2%, 600ng

107590

Twist cfDNA Pan-Cancer Reference Standard v2 VAF 5%, 600ng

*Kit includes 7 VAFs individually: 0% (WT), 0.1%, 0.25%, 0.5%, 1%, 2%, 5% at 300ng each tube

*For research use only. Not for use in any diagnostic or clinical procedures.

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