Skip to main content

Advertisement

Table 1 Specimen requirements and assay specifications of KRAS genotyping by laboratories

From: Reliability of KRAS mutation testing in metastatic colorectal cancer patients across five laboratories

  Lab #1 (Sequencing) Lab #2 (Sequencing) Lab #3 (Sequencing) Lab #4 (Primer Extension) Lab #5 (Real Time PCR)
Specimen Requirements Preferred sample type*: Slides from FFPE block 1  H&E stained slide sections with tumor circled; 4 matching unstained slides, 10 microns each. Preferred sample type: Archival FFPE or frozen surgical biopsies confirmed to contain >50% tumor by a surgical pathologist. 1 H&E slide; 5 unstained sections, 10 microns each. Preferred sample type: FFPE tissue 6 unstained sections, 10 microns each. Preferred sample type: Pre-cut slides from FFPE. Send all slides within 5–7 days of cutting. Air dry. Do not oven dry. Store specimen at room temperature (20–23.5°C). 5 unstained sections, 7 microns each Preferred sample type: FFPE block, unstained slides, or fresh snap frozen biopsy 5 unstained sections, 7 microns each
Genotyping Method: PCR amplification followed by Direct Sanger sequencing (Big Dye v. 1.1) Detected mutations: KRAS codons 12 and 13 Method: PCR amplification followed by standard bidirectional sequencing on ABI 3100. Detected mutations: KRAS codons 12 and 13 Method: PCR amplification followed by sequencing. Detected mutations: KRAS codons 12, 13 and 61 Method: Single nucleotide primer extension with fragment analysis by capillary electrophoresis using a modified SNaPshot assay. Detected mutations: KRAS codons 12 and 13 Methods are propietary: qualitative real time PCR Detected mutations: KRAS codons 12 and 13
Lower Limit of Detection 20% when ≥ 40% tumor cells present 20% 15-20% 10% when ≥ 2% tumor cells present 1-5%
  1. *For this study, slides prepared from Formalin-fixed paraffin embedded (FFPE) blocks were sent to each lab.