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Poster on effect of wash buffers on bacterial cell surface charge Wenfa Ng 22 August 2017.pdf (450.7 kB)

Zeta Potential of Bacterial cells: Effect of Wash Buffer

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posted on 2017-08-22, 01:58 authored by Wenfa NgWenfa Ng

Bacterial surface charge is determined by the ensemble of specifically and nonspecifically adsorbed ions and molecules. Its measurement via the proxy, zeta potential, defined as the electric charge at the shear plane (a short distance from the cell surface), meant that nonspecific adsorption of ions hampers detection of a realistic estimate of cell surface charge, useful for solving a variety of practical problems. Various wash buffers of differing composition (and ionic strength) are used for removing nonspecifically adsorbed ions. Nevertheless, given myriad wash buffers available, systematic studies evaluating their relative performance is needed, and would yield information useful for guiding buffer choice. Using Escherichia coli DH5α (ATCC 53868) grown in two media with different salt content as experiment model, a study examined the relative efficacy of commonly used wash buffers in removing nonspecifically adsorbed ions via ionic strength-dependent charge screening. Specifically, zeta potential as a function of pH from 1 to 12 was used as readout, where differences in the overall zeta potential-pH profile and point of zero charge (pHzpc) provide insights on the relative effectiveness of different wash buffers. Preliminary results suggested that removal of nonspecifically adsorbed ions was positively correlated with wash buffer ionic strength. More important, an operative window in ionic strength might exist, its limits bounded by a threshold ionic strength (i.e., 0.15M) below which there would be no removal of nonspecifically adsorbed ions, and an upper limit (0.6M) beyond which, intrinsic cell surface cations important in maintaining cell envelope integrity might be removed, leading to significant cell surface changes and erroneous zeta potential values. Collectively, presented results should be useful in serving as guidelines for designing appropriate wash buffers used in preparing bacterial cell samples for zeta potential analysis. Nevertheless, medium-specific secretion of metabolites by bacteria meant that the set of adsorbed molecules differ between medium used; thereby, highlighting the need for optimization of wash buffer composition for each strain and medium investigated.

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