Mapping Surface Charge Distribution of Single-Cell via Charged Nanoparticle

Many bio-functions of cells can be regulated by their surface charge characteristics. Mapping surface charge density in a single cell’s surface is vital to advance the understanding of cell behaviors. This article demonstrates a method of cell surface charge mapping via electrostatic cell–nanoparticle (NP) interactions. Fluorescent nanoparticles (NPs) were used as the marker to investigate single cells’ surface charge distribution. The nanoparticles with opposite charges were electrostatically bonded to the cell surface; a stack of fluorescence distribution on a cell’s surface at a series of vertical distances was imaged and analyzed. By establishing a relationship between fluorescent light intensity and number of nanoparticles, cells’ surface charge distribution was quantified from the fluorescence distribution. Two types of cells, human umbilical vein endothelial cells (HUVECs) and HeLa cells, were tested. From the measured surface charge density of a group of single cells, the average zeta potentials of the two types of cells were obtained, which are in good agreement with the standard electrophoretic light scattering measurement. This method can be used for rapid surface charge mapping of single particles or cells, and can advance cell-surface-charge characterization applications in many biomedical fields.

Mapping Surface Charge Distribution of Single-Cell via Charged Nanoparticle

Background: Many bio-functions of cells can be regulated by their surface charge characteristics. Mapping surface charge density in a single cell’s surface is vital to advance the understanding of cell behaviors. Results: This article demonstrates a method of cell surface charge mapping via electrostatic cell–nanoparticle interactions. Nanoparticles with fluorescence were used as the marker to investigate single cells’ surface charge distribution. The nanoparticles with opposite charges were electrostatically bonded to the cell surface; a stack of fluorescence distribution on a cell’s surface at a series of vertical distances was imaged and analyzed. By establishing a relationship between fluorescence light intensity and surface charge density, cells’ surface charge distribution was quantified from the fluorescence distribution. Two types of cells, HUVECs and Hela cells, were tested. From the measured surface charge density of a group of single cells, the average zeta potential of the two types of cells was obtained, which is in good agreement with the standard electrophoretic light scattering measurement. Conclusions: This method can be used for rapid surface charge mapping of single particles or cells and can advance cell-surface-charge characterization applications in many biomedical fields.

A chip device to determine surface charge properties of confluent cell monolayers by measuring streaming potential

Chip device to monitor streaming potential of confluent cell layers reflecting cell surface charge important for the function of biological barriers.

Evolution of Daptomycin Resistance in Coagulase-Negative Staphylococci Involves Mutations of the Essential Two-Component Regulator WalKR

ABSTRACTCoagulase-negative staphylococci (CoNS) represent one of the major causes of health care- and medical device-associated infections. Emerging antimicrobial resistance has complicated the treatment of systemic infections caused by CoNS. Here, we describe the prevalence of antimicrobial resistance in clinical CoNS strains from a tertiary care hospital over a 4-year period, and we observed a significant increase in resistance to daptomycin. Notably,Staphylococcus capitisaccounted for the majority of these daptomycin-resistant (DAP-R) CoNS. To further investigate the mechanisms of daptomycin resistance in CoNS, daptomycin-susceptible clinical strains ofS. capitisandStaphylococcus epidermidisunderwentin vitrodaptomycin exposure to generate DAP-R CoNS mutants. Unlike that seen withStaphylococcus aureus, alteration of cell surface charge was not observed in the DAP-R CoNS strains, but biofilm formation was compromised. Whole-genome sequencing analysis of the DAP-R CoNS strains identified single nucleotide polymorphisms (SNPs) inwalKR, the essential two-component regulatory system controlling cell wall biogenesis. PCR and sequencing ofwalKandwalRfrom 17 DAP-R CoNS clinical isolates identified seven nonsynonymous mutations. The results were confirmed by the recreation of thewalKSNP inS. epidermidis, which resulted in reduced susceptibility to daptomycin and vancomycin. This study highlights the significance of CoNS in evolving daptomycin resistance and showed thatwalKRis shared among the staphylococcal species and is involved in antibiotic resistance development. Notably, we did not observe mutations in genes responsible for phospholipid biosynthesis or an altered cell surface charge, suggesting that reduced daptomycin susceptibility in CoNS may emerge in a fashion distinct from that inS. aureus.

Zeta potential of Escherichia coli DH5α grown in different growth media

Proteins and metabolites typically adsorb to bacterial cell surface through a variety of mechanisms such as van der Waals attraction and electrostatic interactions, and forms a layer of nonspecifically adsorbed ions and molecules on the cell surface. Thus, the bacterial cell surface charge comprised the contribution from the cell wall as well as layers of nonspecifically adsorbed ions and molecules on the cell surface. This is the cell surface charge perceived by other bacterial cells in the growth medium. Given that different growth medium comprises different ensemble of proteins and metabolites that could adsorb onto the cell surface of bacteria, it is important to examine the effect of growth in different medium on the cell surface characteristics of bacterial cells using zeta potential as the proxy parameter. Defined at the shear plane, zeta potential provides a comprehensive view of the cell surface charge that include the nonspecifically adsorbed ions and molecules on the cell surface and the intrinsic electric charges in the cell wall. Using Escherichia coli DH5α (ATCC 53868) as model organism, experiments performed with deionized water as wash and resuspension buffer revealed that the zeta potential-pH profiles of cells grown in LB Lennox, and LB Lennox with 2 g/L glucose overlapped each other over the entire pH range from 2 to 9. This suggested that there was little physiological adaptation of the cell envelope of cells grown in LB Lennox supplemented with 2 g/L glucose, which indicated that the medium could be used for increasing biomass yield without affecting cell surface characteristics. Similarly, the zeta potential-pH profiles of E. coli DH5α grown in LB Lennox and LB Lennox buffered with 89 mM potassium hydrogen phosphate buffer also overlapped each other, which highlighted that the buffered medium did not elicit physiological adaptation of the cell envelope. However, supplementation of the LB Lennox (buffered) medium with 6 g/L glucose resulted in a more negatively charged zeta potential-pH profile in the pH range from 4 to 12 compared to that during growth in LB Lennox. Growth of E. coli DH5α in other media such as Tryptic Soy Broth (TSB), formulated medium, and formulated medium with 6 g/L glucose also resulted in more negatively charged zeta potential-pH profiles compared to that during growth in LB Lennox medium. However, the point-of-zero-charge (pHzpc) of cells grown in TSB and formulated medium were the same as that of cells grown in LB Lennox medium. Collectively, physiological adaptation to growth in different media as well as different ensemble of proteins and metabolites nonspecifically adsorbed to the cell surface would generally result in different zeta potential-pH profiles of bacteria cultivated in growth media of different compositions. Understanding the cell surface charge characteristics of E. coli DH5α grown in different media would thus help unveil the mysteries of cell-cell interactions in the medium.

Zeta potential of Escherichia coli DH5α grown in different growth media

Proteins and metabolites typically adsorb to bacterial cell surface through a variety of mechanisms such as van der Waals attraction and electrostatic interactions, and forms a layer of nonspecifically adsorbed ions and molecules on the cell surface. Thus, the bacterial cell surface charge comprised the contribution from the cell wall as well as layers of nonspecifically adsorbed ions and molecules on the cell surface. This is the cell surface charge perceived by other bacterial cells in the growth medium. Given that different growth medium comprises different ensemble of proteins and metabolites that could adsorb onto the cell surface of bacteria, it is important to examine the effect of growth in different medium on the cell surface characteristics of bacterial cells using zeta potential as the proxy parameter. Defined at the shear plane, zeta potential provides a comprehensive view of the cell surface charge that include the nonspecifically adsorbed ions and molecules on the cell surface and the intrinsic electric charges in the cell wall. Using Escherichia coli DH5α (ATCC 53868) as model organism, experiments performed with deionized water as wash and resuspension buffer revealed that the zeta potential-pH profiles of cells grown in LB Lennox, and LB Lennox with 2 g/L glucose overlapped each other over the entire pH range from 2 to 9. This suggested that there was little physiological adaptation of the cell envelope of cells grown in LB Lennox supplemented with 2 g/L glucose, which indicated that the medium could be used for increasing biomass yield without affecting cell surface characteristics. Similarly, the zeta potential-pH profiles of E. coli DH5α grown in LB Lennox and LB Lennox buffered with 89 mM potassium hydrogen phosphate buffer also overlapped each other, which highlighted that the buffered medium did not elicit physiological adaptation of the cell envelope. However, supplementation of the LB Lennox (buffered) medium with 6 g/L glucose resulted in a more negatively charged zeta potential-pH profile in the pH range from 4 to 12 compared to that during growth in LB Lennox. Growth of E. coli DH5α in other media such as Tryptic Soy Broth (TSB), formulated medium, and formulated medium with 6 g/L glucose also resulted in more negatively charged zeta potential-pH profiles compared to that during growth in LB Lennox medium. However, the point-of-zero-charge (pHzpc) of cells grown in TSB and formulated medium were the same as that of cells grown in LB Lennox medium. Collectively, physiological adaptation to growth in different media as well as different ensemble of proteins and metabolites nonspecifically adsorbed to the cell surface would generally result in different zeta potential-pH profiles of bacteria cultivated in growth media of different compositions. Understanding the cell surface charge characteristics of E. coli DH5α grown in different media would thus help unveil the mysteries of cell-cell interactions in the medium.