Fei tecnai g2 biotwin

Morphological changes of E. coli O157 strain EDL933 following incubation with hGRNL, bNK2A, or pNKL peptides were examined by TEM. Overnight cultures of O157 strain EDL933 were centrifuged (5000 rpm for 30 min) and diluted to ~2×10⁹ CFU/mL in NaPB. Fifty microliters of bacterial suspension were placed in 96-well flat-bottom polystyrene plates (Falcon #351172) and hGRNL, bNK2A, pNKL (20 μM final concentration) or NaPB were added and incubated at 37˚C for 15 (or 30 min) with continuous shaking. The bacterial suspensions were mixed with 120 μL of 3% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.4) and processed for TEM as described previously [26 (link)]. Ultrathin sections were stained with uranyl acetate and lead citrate and, examined with a TEM (FEI Tecnai G2 Biotwin; FEI Co., Hillsboro, OR) and images were captured with Advanced Microscopy Technologies (AMT Inc., Danvers, MN) imaging camera. The final figures were prepared using Adobe Photoshop Elements 11 (Adobe Systems Inc., San Jose, CA).

Morphological changes of bacteria following incubation with NK2A and Sc-NK2A peptides were studied by TEM. An overnight culture of Salmonella (isolate SX238) was centrifuged and resuspended in PBS. One hundred microliters of bacterial suspension were placed in a Honeycomb plate (~ 1 × 10⁷ CFU/well) and NK2A, Sc-NK2A (20 µM), and PBS were added and incubated at 37 °C with continuous shaking for 1 h. The bacterial suspensions were mixed with an equal volume of 3% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.4) and processed for TEM as described before^{24 (link)}. Sections were examined with a FEI Tecnai G2 Biotwin (FEI Co., Hillboro, OR) TEM and images were taken with Advanced Microscopy Technologies (AMT Inc., Danvers, MN) imaging camera.

Approximately, 1 × 10⁸ CFUs of M. bovis in 100 μl of PBS was transferred to a non-tissue culture treated flat-bottom 96-well plate and incubated with 30 μM of NK-lysin peptides or PBS at 37°C in a humidified atmosphere of 5% CO₂ for 60 min. The bacterial suspension was mixed with an equal volume of 3% glutaraldehyde in 0.1 M cacodylate buffer, pH 7.4 and processed for transmission electron microscopy as described previously [17 (link), 28 (link)]. Briefly, after fixation, bacterial pellets were rinsed in cacodylate buffer, post-fixed in 1% osmium tetroxide, dehydrated in alcohols, and embedded in epoxy resin. Ultrathin sections of the bacterial pellets were cut and stained with uranyl acetate and lead citrate. Sections were examined with a FEI Tecnai G2 Biotwin (FEI Co., Hillboro, OR) transmission electron microscope and images were taken with Advanced Microscopy Technologies (AMT Inc., Danvers, MN) imaging camera.