The effects of cheopin on the integrity of the bacterial outer membranes were studied by measuring the β-lactamase activity as previously described73 (link) with slight modifications. Briefly, Y. pestis strains carrying the β-lactamase-encoding plasmid pUC19 were grown in the presence of ampicillin to an O.D. of 0.2–0.3, washed twice with 10 mL of PBS-CM (pH 7.4, supplemented with 1 mM CaCl2 and 0.5 mM MgCl2) by centrifugation (5 min, 5000 × g) and resuspended (1×107 CFU/mL) in PBS-CM containing 180 μM CENTA, a chromogenic β-lactamase substrate (Sigma). Cells (100 μL) were then incubated with 4 μM cheopin or Fast Break lysis solution (Promega, Madison, WI) and absorbance at 405 nm was measured for 1h using a BioTek Synergy H1M plate reader set to 28°C.
Fast break lysis solution
Manufactured by Promega
Fast Break lysis solution is a laboratory product designed for the rapid lysis of cells. It facilitates the extraction and release of cellular contents, including proteins, nucleic acids, and other biomolecules, from a variety of cell types. The solution is formulated to provide efficient and consistent lysis without the need for extensive sample preparation or disruption methods.
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2 protocols using fast break lysis solution
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Cheopin Impacts Bacterial Membrane Integrity
2
Cheopin Membrane Integrity Assay
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The effects of cheopin on the integrity of the bacterial outer membranes were studied by measuring the β-lactamase activity as previously described [59] Cells (100 µL) were then incubated with 4 µM cheopin or Fast Break lysis solution (Promega, Madison, WI) and absorbance at 405 nm was measured for 1h using a BioTek Synergy H1M plate reader set to 28°C.
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