Bacteria were obtained from American Type Culture Collection (ATCC; Manassas, VA, USA) as listed in Table S2 (see ESI). Bacteria were cultured in Nutrient broth (NB; Bacto Laboratories, catalog No. 234000) or Muller-Hinton broth (MHB; Bacto Laboratories, catalog No. 211443) at 37 °C overnight with shaking (~180 RPM). A sample of each culture was diluted 50-fold in fresh MHB and incubated at 37 °C for 1.5–3 h with shaking (~180 RPM). Compound stock solutions were prepared as 10 mg/mL in DMSO and colistin was dissolved in milli-Q water at 5.12 mg/mL. The compounds, at twice the final desired concentration, were serially diluted 2-fold across the wells of 96–well plates (Non-Binding Surface, Corning, catalog No. 3641). Mid-log phase bacterial cultures (after 1.5–3 h incubation) were diluted to a final concentration of 5 × 105 colony forming units (CFU)/mL, and 50 µL was added to each well giving a final compound concentration range of 32 µg/mL to 0.015 µg/mL (DMSO ≤ 1%). MICs were determined visually after 20 h of incubation at 37 °C, with the MIC defined as the lowest compound concentration at which no bacterial growth was visible.
Non binding surface
Manufactured by Corning
Non-Binding Surface is a specialized lab equipment product from Corning. It is designed to provide a surface that minimizes the binding or attachment of materials, enabling efficient sample handling and processing. The core function of this product is to maintain sample integrity and facilitate easy separation or release of materials during laboratory procedures.
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Lab products found in correlation
4 protocols using non binding surface
1
Antimicrobial Susceptibility Testing Protocol
2
High-Throughput smFCS Analysis of RNA-Protein Interactions
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smFCS experiments were scaled up into 96 well plates. Two 96 well plates (Non-Binding Surface, Corning) were used. PS1 RNA was labelled and gel purified as described earlier and HBV dimer was purified as described above. Each well contained 200 μL of 15 nM PS1 in re-assembly buffer. As in smFCS, ten 2 μL injections of 2.5 μM dimer in dissociation buffer were performed. A Perkin-Elmer Envision plate reader was used to carry out the injections and record the anisotropy of the PS1 RNA (FITC excitation and emission filters). VLPs were purified away from free RNA and capsid using a 1.33 g/mL caesium chloride gradient and spun at 113,652 x g for 90 hours using an SW40Ti rotor. A single band was observed and fractionated. The band was dialysed into reassembly buffer to remove caesium chloride. The 2 mL fraction of VLP was concentrated to 200 μL using an Amicon 100 kDa MWCO spin concentrator.
3
High-Throughput smFCS Analysis of RNA-Protein Interactions
4
Turbidity Assay of Protein Samples
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Protein samples were prepared in 50 mM Tris-HCl buffer, 40 mM NaCl, and 1 mM DTT pH 7.0 or 50 mM Tris-HCl buffer, 150 mM NaCl, 1 mM DTT, and 10% PEG, pH 7.0, in a total volume of 100 μl. The samples were incubated at room temperature for 15 min, and absorbance at 370 nm was measured using a Beckman Coulter DTX 880 multimode detector. All the conditions were measured in 96-well plates (Corning nonbinding surface). Each turbidity assay was reproduced in triplicate.
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