High throughput platforms such as Next generation sequencing (NGS) and quantitative PCR (qPCR) platforms give researchers much greater power when profiling cfDNA samples. However, the use of these presents challenges surrounding sample variability, platform bias, and potential failure to detect biomarkers. Horizon’s cfDNA products are all derived from human cell lines, and are fragmented to an average size of 160 bp to closely resemble cfDNA extracted from human plasma.

This format ensures that they may applied to a wide-range of cfDNA NGS and qPCR assays including high-throughput platforms such as NGS and qPCR give researchers greater power when profiling cfDNA samples. Horizon has developed the cfDNA Reference Standard range to support the development and continued validation of cfDNA assays.

The Multiplex I cfDNA Reference Standard Set covers multiple engineered single nucleotide variants (SNVs/SNPs) with 8 mutations at 5%, 1%, and 0.1% allelic frequencies. Batch-specific copy number values measured via ddPCR are provided with each Set.

With this product you are able to:

• Analyze the sensitivity and specificity of your assay
• Gain certainty of the limit of detection and limit of quantification
• Optimize and validate new cancer panels and routinely monitor the performance of your assay

Technical data

Format: cfDNA

Genes covered: EGFR, KRAS, NRAS, PIK3CA

Cosmics: 1,500+

Allelic frequencies: 5%, 1%, 0.1% and 0% (100% wild type)

Buffer: Tris-EDTA (10mM Tris-HCl, 1mM EDTA), pH 8.0

Product information

Fragment size: 160 bp

Unit size: 350 ng per vial

Concentration: 20 ng/µl

Verified mutations: The Multiplex cfDNA Reference Standard has been manufactured using multiple engineered cell lines and mixed to generate multiple Allelic Frequencies in multiplex. Two of the four products in this set (HD776 and HD777) have been verified by whole exome sequencing. The NGS determined allelic frequency and read depths for HD777 are reported in the table below.

a – Not detected - did not meet detection threshold, below LOD
b – 0.00% expected VAF, not detected by variant caller

General information

Storage: 4°C

Expiration: See all product shelf life information

Quality control

Fragmentation size: D1000 DNA ScreenTape assay

Allelic frequency: Droplet Digital™ PCR

Quantification: Qubit dsDNA BR Assay (post-fragmentation)