Genomic DNA was isolated from the provided tumor specimen using the cobas® cfDNA Sample Preparation Kit for plasma. Mutation detection is achieved through real-time PCR analysis on cobas® z480 analyzer. The cobas® EGFR Mutation Test v2 detects the following mutations: exon 18 (G719X), exon 19 (deletions and complex mutations), exon 20 (S768I, T790M, insertions) and exon 21 (L858R, L861Q).
Non-Small-Cell Lung Cancer (NSCLC)
Plasma (K2EDTA), FROZEN
4 mL frozen plasma from K2EDTA tubes
BD K2EDTA 4 mL blood collection tubes. Collect three tubes of whole blood in accordance with the collection instructions specified below.
It is recommended that patients be referred to a LabCorp Patient Service Center for collection. Otherwise, take care to follow the procedure of specimen collection and preparation outlined below:
Blood should be collected according to your laboratory’s procedure for venipuncture using ONLY BD K2EDTA 4 mL blood collection tubes. Plasma preparation should be performed within 4 hours after blood is collected. Store, transport and ship plasma at frozen condition only. The plasma sample can be stored frozen, at <-70°C for up to 12 months.
Note: Do not freeze whole blood samples.
Plasma Specimen Preparation
• Immediately following specimen collection, label all tubes with appropriate patient identification information (two patient identifiers, minimum and label "plasma" on transport tube).
• The brake setting of the Drucker 642e at LabCorp service center is at 3 or 4 (manufacture setting).
• Centrifuge the three, BD K2EDTA blood collection tubes for 10 min at 1600 ± 90 rcf. [For conversion of RPM (revolutions per minute) to rcf (relative centrifugal force), refer to the centrifuge manufacture’s user manual]
• Remove blood collection tubes from the centrifuge (plasma sample will be rejected if it is hemolyzed).
• Using a fresh 6 inch disposable transfer pipette, transfer plasma from three, 4 mL blood collection tubes to one 8.5 mL Sarstedt centrifuge tube (55.598.006). One, 8.5 mL tubes will be collected from three, 4 mL blood collection tubes.
• Centrifuge plasma in the 8.5 mL centrifuge tube for 10 min at 1600 ± 90 rcf.
• Using a fresh, 6 inch disposable transfer pipette or serological pipette, transfer 4 mL of plasma from one 8.5 mL centrifuge tube into a labeled 7.0 mL Sarstedt screw cap, flat bottom purple frozen transport tube (62.550.019). One, 7.0 mL tube will be collected.
• Ship and store frozen plasma at <-70°C. Stable for up to 12 months.
Note: Take care not to disturb or transfer the buffy coat (white blood cells) layered above the red blood cells in the blood collection tube after the first centrifugation or sedimented at the bottom of the centrifuge tube after the second centrifugation.
Causes for Rejection
Hemolyzed blood; Plasma not frozen; Plasma volume <2mL per tube. Leaking or broken tubes.
Frozen; The plasma sample could be stored frozen, at <-70°C for up to 12 months.
The FDA-approved cobas® EGFR mutation test v2 is a real-time PCR test for the qualitative detection of defined mutation in exon 18, 19, 20 and 21 of the EGFR gene in non−small-cell lung cancer (NSCLC) patients. The FDA-approved plasma test is indicated as a companion diagnostic to aid in identifying NSCLC patients for treatment with the targeted therapies erlotinib (TARCEVA®) or osimertinib (TAGRISSOTM).
- TARCEVA – Exon 19 deletions and L858R mutations
- TAGARISSO – T790M mutations
Detection of mutation is depended on sample integrity and the amount of amplifiable DNA present in the specimen. The methods used in this assay are highly selective and, depending on the total amount of DNA present, can detect mutations with ≤100 copies of mutant DNA per mL of plasma using the standard imput of 25uL of DNA stock per reaction well.
epidermal growth factor receptor
1. Cobas® EGFR Mutation Test v2 (IVD) [package insert]. Indianapolis, Ind: Roche; Nov 2015.
2. Jänne PA, Engelman JA, Johnson BE. Epidermal growth factor receptor mutations in non-small-cell lung cancer: Implications for treatment and tumor biology. J Clin Oncol. 2005 May 10; 23(14): 3227-3234. PubMed 15886310