KRAS mutation testing in metastatic colorectal cancer

Table 2 Methods used for KRAS mutation testing[45,55,60-62]

Method	Sensitivity (mutant/wild-type) (%)	Turnaround time	Main advantages	Main disadvantages
Sanger sequencing	20–30	Slow (4 d to 2 wk)	Detects all possible mutations, cost-effective	Insensitive, time consuming, open PCR system is easily contaminated
Pyrosequencing	5	Rapid	Detects all possible mutations, sensitive	Open PCR system is easily contaminated
Real-time PCR with HRMA	5	Rapid	Rapid, closed PCR system, detects all possible mutations (heterozygous and homozygous)	Occasionally difficult to distinguish between mutation types
Allele-specific real-time PCR	10	Rapid	Rapid, closed PCR system	Detects only the 7 most common mutations, requires more tissue for analysis compared with other methods
RFLP with sequencing	0.1	Slow (4 d to 2 wk)	Sensitive	Requires confirmation by sequencing, complicated
DxS (ARMS/S)	1	Rapid	Sensitive, time-saving	Expensive, detects specific mutations targeted by the designed primers
COLD-PCR with sequencing	1-2.5	Rapid	Sensitive, cost-effective, detects all possible mutations	-

ARMS: Amplification refractory mutation system; RFLP: Restriction fragment length polymorphism; PCR: Polymerase chain reaction; COLD-PCR: Coamplification at lower denaturation temperature PCR.