BACKGROUND: KRAS, BRAF and PIK3CA mutations are frequently observed in 
colorectal cancer (CRC). In particular, KRAS mutations are strong predictors for 
clinical outcomes of EGFR-targeted treatments such as cetuximab and panitumumab 
in metastatic colorectal cancer (mCRC). For mutation analysis, the current 
methods are time-consuming, and not readily available to all oncologists and 
pathologists. We have developed a novel, simple, sensitive and fully automated 
molecular diagnostic system (AMDS) for point of care testing (POCT). Here we 
report the results of a comparison study between AMDS and direct sequencing (DS) 
in the detection of KRAS, BRAF and PI3KCA somatic mutations.
METHODOLOGY/PRINCIPAL FINDING: DNA was extracted from a slice of either frozen 
(n = 89) or formalin-fixed and paraffin-embedded (FFPE) CRC tissue (n = 70), and 
then used for mutation analysis by AMDS and DS. All mutations (n = 41 among 
frozen and 27 among FFPE samples) detected by DS were also successfully (100%) 
detected by the AMDS. However, 8 frozen and 6 FFPE samples detected as wild-type 
in the DS analysis were shown as mutants in the AMDS analysis. By 
cloning-sequencing assays, these discordant samples were confirmed as true 
mutants. One sample had simultaneous "hot spot" mutations of KRAS and PIK3CA, 
and cloning assay comfirmed that E542K and E545K were not on the same allele. 
Genotyping call rates for DS were 100.0% (89/89) and 74.3% (52/70) in frozen and 
FFPE samples, respectively, for the first attempt; whereas that of AMDS was 
100.0% for both sample sets. For automated DNA extraction and mutation detection 
by AMDS, frozen tissues (n = 41) were successfully detected all mutations within 
70 minutes.
CONCLUSIONS/SIGNIFICANCE: AMDS has superior sensitivity and accuracy over DS, 
and is much easier to execute than conventional labor intensive manual mutation 
analysis. AMDS has great potential for POCT equipment for mutation analysis.