High speed microcentrifuge

A commercial food grade lactic acid (88% v/v) was obtained from Xena Inc (Xena International Inc; Polo IL, USA) and lactic acid solutions were prepared in TSB at concentrations of 0.5, 1, 1.5, 2, 2.5, and 5% v/v. The pH of lactic acid was determined using a pH meter (Oakton pH 510 Benchtop Meter, Oakton Instruments, Vernon Hills, IL, USA) with a sensitivity of 0.01 and 2-point calibration. The concentration at which lactic acid could prevent survival and regrowth of the STEC strains was considered as the bactericidal concentration. The bactericidal concentration of lactic acid for all the bacterial strains was evaluated as follows: Each 900 μl of 0.5, 1, 1.5, 2, 2.5, and 5% v/v lactic acid was inoculated with 100 μl of bacterial culture (6.89 ± 0.72 log CFU/ml) for 300 s. Solution was centrifuged immediately after exposure for 1 min at 13000 × g using a Corning high speed microcentrifuge (Corning Inc., Corning, NY, USA). The supernatant was discarded, and pellets were resuspended in 1 ml sterile deionized water (SDW). From the resuspended solution, 100 μl was transferred to 100 μl of 2 × TSB in 96-well plates (Costar^® 96 Well Flat Bottom, Corning Life Sciences Inc. ME, USA) and incubated for 24 h at 37°C. The plates were observed for turbidity to determine survival and regrowth after incubation by determining the OD_{600 nm} using the BioTek Cytation 3 multi-mode plate reader.

A commercial food grade lactic acid (88% v/v) was obtained from Xena Inc (Xena International Inc; Polo IL, USA) and lactic acid solutions were prepared in TSB at concentrations of 0.5, 1, 1.5, 2, 2.5, and 5% v/v. The pH of lactic acid was determined using a pH meter (Oakton pH 510 Benchtop Meter, Oakton Instruments, Vernon Hills, IL, USA) with a sensitivity of 0.01 and two-point calibration. The concentration at which lactic acid could prevent survival and regrowth of the STEC strains was considered as the bactericidal concentration. The bactericidal concentration of lactic acid for all the bacterial strains was evaluated as follows: Each 900 μl of 0.5, 1, 1.5, 2, 2.5, and 5% v/v lactic acid was inoculated with 100 μl of the bacteria for 300 s. Solution was centrifuged immediately after exposure for 1 min at 13000 × g using a Corning high speed microcentrifuge (Corning Inc., Corning, NY, USA). The supernatant was discarded, and pellets were resuspended in 1 ml sterile deionized water (SDW). From the resuspended solution, 100 μl was transferred to 100 μl of 2 × TSB in 96-well plates (Costar^® 96 Well Flat Bottom, Corning Life Sciences Inc. ME, USA) and incubated for 24 h at 37 °C. The plates were observed for turbidity to determine survival and regrowth after incubation by determining the OD_{600 nm} using the BioTek Cytation multi-mode plate reader (Figure 1).