Pre-concentration and separation of bacteria by volume coupling electrophoresis on supercritical water-etched fused silica capillary with two segments of different internal diameters and inner surface roughnesses

Abstract

The transient isotachophoretic stacking and sweeping was used for the on-line large-volume sample pre-concentration of bacteria, Escherichia coli and Staphylococcus aureus cells (methicillin-susceptible or methicillin-resistant), in the initial stage of micellar electrokinetic chromatography using a non-ionogenic surfactant or of capillary electrophoresis, respectively. These procedures were employed in single-piece fused silica capillary etched with supercritical water with two different internal diameter segments featuring different inner surface roughness. Large volumes (maximum 2.8 μL) of the high conductivity sample matrices, physiological saline solution, urine or blood (with purification step), spiked with examined cells were injected into the wider end of a capillary with an inlet inner diameter 195 μm. This novel on-line combination of preconcentration strategies for cells produced an up to 680-fold increase in sensitivity for E. coli or S. aureus cells. The average calculated resolutions, R, for five selected methicillin-susceptible or methicillin-resistant strains were found to be 6.3 for the agar-cultivated and 14.9 for the blood-incubated cells. A low number of bacteria similar to those in clinical samples were also tested. The modified surface roughness step helped to significantly narrow the cell zones and to increase resolution. The migration velocities of E. coli agar-cultivated and blood-incubated cells were approximately the same as those of S. aureus, probably due to the minimal differences in their surface properties. This procedure, on-line pre-concentration and separation of bacteria, is rapid and provides good reproducibility and repeatability.

Recent advances in enhancing the sensitivity of electrophoresis and electrochromatography in capillaries and microchips (2016–2018)

“…To combine sweeping and tITP, a BGE containing a pseudostationary phase is used and the sample is devoid of this phase, with typically the ITP stage occurring prior to the psuedophase sweeping and separating the ITP‐stacked analytes. tITP‐sweeping was applied for the analysis of proteins in artificial urine, bacteria in urine and blood , and seven hydrophobic chlorophenol residues in wines samples .…”

Recent progress in analytical capillary isotachophoresis

“…With UV detection at 270 nm, this method was successfully applied also to other types of cells (BGE = TE: 40 mM Tris, 275 mM glycine, 5.0 mM NaOH, 2.5 mM Mg(OH) 2 with 0.0125% polyethylene oxide, pH 8.6; LE: Cl − from sample). Horká and co‐workers used tITP stacking and sweeping for the sample preconcentration of E. coli and Staphylococcus aureus methicillin‐susceptible or methicillin‐resistant bacteria cells in spiked urea and blood. This novel online combination of preconcentration strategies for cells produced up to 680‐fold increase in sensitivity for E. coli or S. aureus cells (BGE = TE: 5 mM phosphate buffer, 5% ethanol, and 0.3% Brij 35, pH 7; LE: Cl − present in the sample, UV detection at 280 nm).…”

Identification of Aspergillus Conidia in Bronchoalveolar Lavage Using Offline Combination of Capillary Electrophoresis in Supercritical Water-Treated Fused Silica Capillary and Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

“…Recently, fused silica (FS) capillaries were etched with supercritical water (SCW) to modify their inner diameter , or to roughen their inner surface for the purposes of electrophoretic separations. − The rough surface is strongly polar and allows adsorption of ionic substances including amphoteric microorganisms . Although most CE separations are performed in smooth FS capillaries, the modification of their inner surface can significantly enhance the separation process. ,,, In addition, the composition of background electrolyte (BGE) in CE analysis must reflect the surface characteristics of the analyzed microorganisms. Highly hydrophobic conidia are an example of sample the analysis of which is quite challenging .…”

“…47 An FS capillary with a roughened part can be advantageously used for dynamic adhesion and subsequent online desorption, concentration, and analysis of microorganisms using a combination of polymer-enhanced transient isotachophoresis (tITP) 36 and MEKC when the charged microbial cells are swept using nonionogenic surfactant Brij 35. 35,36,43,48,49 The best results were achieved when the roughened part was located at the anodic side of the capillary. 36,39,41,43 The aim of this study was to develop a reliable method that would contribute to rapid diagnosis of Aspergillus infection and, consequently, to timely initiation of therapy.…”

“…35,36,43,48,49 The best results were achieved when the roughened part was located at the anodic side of the capillary. 36,39,41,43 The aim of this study was to develop a reliable method that would contribute to rapid diagnosis of Aspergillus infection and, consequently, to timely initiation of therapy. The method should ideally allow direct analysis of clinical specimen without need of previous cultuvation or complicated sample treatment.…”