VarTrace EGFR Mutation Assay

For research use only. Not for use in diagnostic procedures.

EGFR mutations are present in 10-30% non-small cell lung cancer (NSCLC) patients. Specific mutations can guide treatment with targeted therapies such as TARCEVA® (erlotinib), TAGRISSO® (osimertinib), IRESSA® (gefitinib).

The VarTrace EGFR Assay enables highly sensitive detection and quantification of EGFR mutations for patients with advanced or metastatic NSCLC. The kit uses NuProbe’s PCR-based Blocker Displacement Amplification (BDA) technology to enable the selective amplification of low abundant sequence variants (SNV and indels) in a background of wildtype DNA. Following PCR enrichment, Sanger sequencing is applied to reveal the identity of the variants.

Key Features

Sensitive

Limit of detection down to 0.1% VAF

Quantitative

Able to quantify mutation VAF in original sample

Versatile

Applicable to a variety of sample types

Comprehensive

Wide range of mutations covered

User-friendly

Easy workflow

Benefits

  • Detects 40 EGFR mutations including G719X substitution mutations in exon 18, deletion mutations in exon 19, T790M, C797S and S768I substitution mutations in exon 20, insertion mutations in exon 20, and L858R and L861Q substitution mutations in exon 21
  • Enables distinction of cis/trans mutations in C797S/T790M
  • Compatible with plasma samples, tumor biopsies, paraffin-embedded sections (FFPE), and fresh frozen tumors
  • Requires 10-100 ng cfDNA from blood plasma, or 10-1000 ng DNA from tissue biopsy
  • Delivers results in 8 hours, with 2.5 hours of hands-on time

Sample Data

  • Variant verification by Sanger sequencing
  • Because NuProbe’s BDA technology enriches the variant allele frequency (VAF) of mutations by over 200-fold, Sanger’s limit of detection of 20% VAF is effectively improved to 0.1% VAF. Sanger sequencing can be applied to confirm the identity of quantities detected and quantitated by qPCR using standard sequencing protocols.
  • Variant verification by Sanger sequencing
  • Because NuProbe’s BDA technology enriches the variant allele frequency (VAF) of mutations by over 200-fold, Sanger’s limit of detection of 20% VAF is effectively improved to 0.1% VAF. Sanger sequencing can be applied to confirm the identity of quantities detected and quantitated by qPCR using standard sequencing protocols.

Workflow

Step 1: DNA Extraction

DNA is extracted from clinical samples with commercial extraction kits such as cobas® cfDNA Sample Preparation Kit, QIAamp® DNA Blood Mini Kit, GeneRead® DNA FFPE Kit.
40 min (40 min hands-on)

Step 2: qPCR BDA Reaction

NuProbe’s qPCR/Sanger kit reagents and clinical samples are prepared for qPCR reactions. After running the reaction, Ct results and BDA products are obtained.
1.5 hrs (30 min hands-on)

Step 3: Sanger Sequencing

BDA products are prepared for Sanger sequencing, including PCR product purification, cycle sequencing and sequencing clean-up.
3.5 hrs (1hr hands-on)

Purified Sanger products are analyzed by capillary electrophoresis.
2 hrs (15 min hands-on)

Compatible Instruments

qPCR

Sanger sequencing