KRAS StripAssay

The Viennalab KRAS XL StripAssay detects mutations in the KRAS gene in patients with metastatic colorectal cancer, in order to establish whether anti-EGFR therapy can be effective.

Why KRAS matters for anti-EGFR therapy

Monoclonal antibodies against the epidermal growth factor receptor (EGFR) block signalling through that receptor. KRAS lies downstream of EGFR in the MAPK signalling pathway. A mutation in KRAS activates the pathway permanently, which cancels the inhibitory effect of the antibody. Genetic screening for these mutations therefore determines whether a patient is eligible for cetuximab or panitumumab.

Mutations covered

The assay covers 29 mutations, distributed over codon 12 (8 mutations), codon 13 (6), codon 59 (3), codon 60 (1), codon 61 (5), codon 117 (3) and codon 146 (3).

Sample material

The assay is used on DNA from formalin-fixed paraffin-embedded (FFPE) tissue among other sources. Note that DNA from FFPE tissue cannot be reliably quantified by UV photometry: measurement at 260 nm substantially overestimates the amount of usable template, so a fluorometric method is required for quantification.

Method

The procedure has three steps: isolation of DNA, PCR amplification of the KRAS region, and hybridisation of the product to probes immobilised on a test strip, followed by a colour reaction. The kit contains the amplification mix, Taq DNA polymerase and dilution buffer, DNAT, typing trays, 20 test strips, hybridisation buffer, wash solutions, conjugate solution, colour developer and a collector sheet.

• Amplification Mix (500 µl)
• Taq Dilution Buffer (500 µl)
• Taq DNA Polymerase (75 U)
• DNAT (1.5 ml)
• Typing Trays (3)
• Teststrips (20)
• Hybridization Buffer (25 ml)
• Wash Solution A (80 ml)
• Wash Solution B (80 ml)
• Conjugate Solution (25 ml)
• Color Developer (25 ml)
• Collector™ Sheet