Kishan Rijal

A method for DNA-based detection of E. coli O157:H7 in a proteinous background using piezoelectric-excited cantilever sensors

We show for the first time the detection of ~700 E. coli O157:H7 (EC) in both buffer and proteinous sample (beef wash) using the virulent gene stx2 as a sensing molecule immobilized on piezoelectric-excited millimeter cantilever (PEMC) sensors without a sample preparation step. Genomic DNA of EC suspended in buffer or beef wash was extracted using a 30-minute procedure, or using a commercial extraction kit, and then was exposed to the sensor immobilized with a 19-mer probe that is complement to stx2 gene. As hybridization occurred, the resonant frequency of the cantilever sensor decreased due to the increased attached mass indicating the presence of stx2 gene in the sample. Wild strain JM101 subjected to the same preparation and procedure did not induce a hybridization response, nor did the genomic extract of EC with a bare sensor. PEMC sensor responses to diluted genomic extracts indicate that a much lower concentration of 700 cells is detectable. In order to compare the results with antibody-based detection, samples with EC at 2500 cells per mL were exposed to antibody-immobilized PEMC sensors which gave reproducible responses confirming previous experiments with such samples.