Molecular diagnostics of human cancers may increase accuracy in prognosis, 
facilitate the selection of the optimal therapeutic regimen, improve patient 
outcome, reduce costs of treatment and favour development of personalized 
approaches to patient care. Moreover sensitivity and specificity are fundamental 
characteristics of any diagnostic method. We developed a highly sensitive 
microarray for the detection of common KRAS and BRAF oncogenic mutations. In 
colorectal cancer, KRAS and BRAF mutations have been shown to identify a cluster 
of patients that does not respond to anti-EGFR therapies; the identification of 
these mutations is therefore clinically extremely important. To verify the 
technical characteristics of the microarray system for the correct 
identification of the KRAS mutational status at the two hotspot codons 12 and 13 
and of the BRAF(V600E) mutation in colorectal tumor, we selected 75 samples 
previously characterized by conventional and CO-amplification at Lower 
Denaturation temperature-PCR (COLD-PCR) followed by High Resolution Melting 
analysis and direct sequencing. Among these samples, 60 were collected during 
surgery and immediately steeped in RNAlater while the 15 remainders were 
formalin-fixed and paraffin-embedded (FFPE) tissues. The detection limit of the 
proposed method was different for the 7 KRAS mutations tested and for the V600E 
BRAF mutation. In particular, the microarray system has been able to detect a 
minimum of about 0.01% of mutated alleles in a background of wild-type DNA. A 
blind validation displayed complete concordance of results. The excellent 
agreement of the results showed that the new microarray substrate is highly 
specific in assigning the correct genotype without any enrichment strategy.