Electrochemical attachment of motile bacterial cells to gold

Selective attachment of Escherichia coli K-12 bacterial cells to charged gold surfaces was demonstrated. Electrostatic binding of E. coli K-12 bacterial cells to positively charged surfaces was observed starting at +750 mV. The binding of E. coli K-12 cells to positively charged gold surfaces is proposed to occur due to long-range electrostatic interactions between the negatively charged O-chain of lipopolysaccharide (LPS) molecules protruding the bacterial cell body and the electrode surface. Removing LPS alters the cellular surface charge and results in cellular attachment to negatively charged surfaces. Thus, applying an electrical potential allows for the direct, real time detection of live, dead or damaged bacterial cells. The attachment of E. coli K-12 bacterial cells to surfaces with an applied potential substantiates the hypothesis that an electrostatic interaction is responsible for the binding of bacterial cells to positively charged molecular assemblies on surfaces used for building bacterial microarrays.     

Attachment of bacterial cells to carbon electrodes

Anodic stripping method was applied to analyze the process of bacterial attachment to the surface of carbon-paste electrodes (CPE). The electrode was immersed for various times in a bacterial cell suspension to allow the cells to attach to its surface. The number of bacterial cells attached to the electrode surface increased along with time. On the other hand, the current derived from the oxidation of a dye, Hoechst, which was adsorbed to the surface after attaching the bacterial cells, decreased along with time. It was considered that the current output, correlated with the amount of dye, adsorbed onto regions where no bacterial cell attached. These results indicate that the bacterial-attachment process can be analyzed by measuring the electric current derived from the dye instead of counting the number of attached cells.     

Using ratchets and sorters to fractionate motile cells of Escherichia coli by length

This paper describes the fabrication of a composite agar/PDMS device for enriching short cells in a population of motile Escherichia coli. The device incorporated ratcheting microchannels, which directed the motion of swimming cells of E. coli through the device, and three sorting junctions, which isolated successively shorter populations of bacteria. The ratcheting microchannels guided cells through the device with an average rate of displacement of (32 +/- 9) microm s(-1). Within the device, the average length of the cells decreased from 3.8 microm (Coefficient of Variation, CV: 21%) at the entrance, to 3.4 microm (CV: 16%) after the first sorting junction, to 3.2 mum (CV: 19%) after the second sorting junction, to 3.0 mum (CV: 19%) after the third sorting junction.     

Screening of bacterial genes responsible for resistance to beta-lactam antibiotics using microarrays with enzymatic detection

The method of hybridization analysis on microarrays with enzymatic detection based on horseradish peroxidase is applied to screen infectious agents of nosocomial and community-acquired infections for beta-lactamase genes causing resistance to beta-lactam antibiotics. The advantages of using this method for the rapid identification of genes are demonstrated. Similarities and differences in the distribution of beta-lactamase genes identified in the infectious agents of nosocomial and community-acquired infections are revealed. The most common type of extended-spectrum beta-lactamases (ESBLs) is CTX-M. The high prevalence of extended-spectrum beta-lactamases, particularly of the TEM-1 beta-lactamase, is demonstrated. Individually or in combination with genes of TEM-1 and SVH-1 beta-lactamases, the genes of subgroup CTX-M-1 beta-lactamases were the most frequently identified in community-acquired infectious agents. There were no cases of the simultaneous detection of multiple ESBLs in community-acquired infectious agents. Much more varied combinations of beta-lactamases were identified in nosocomial infectious agents: a combination of extended-spectrum beta-lactamases and broad-spectrum beta-lactamases was identified in 62% of strains and the simultaneous presence of two different types of ESBLs was identified in 18% of strains.     

Attachment of drugs to polydimethylsiloxanes

Polydimethylsiloxane, which is a biologically inert polymer, was used as a carrier polymer for drugs. Functional groups were introduced into the polymer to enable attachment to drugs. Polymers (and oligomers) having carboxyl, amine or hydroxyl groups were prepared from polydimethylsiloxanes having different distributions of SiH groups which were added to the C=C double bond of unsaturated acids, amines or alcohols. These polymers were reacted with suitable functional groups on drugs such as digoxin, quinidine, barbiturates, amphetamine, naloxone and atropine.     

Dissociative Electron Attachment to Hexachlorobenzene

Gas phase molecules of hexachlorobenzene (C₆Cl₆) were investigated by means of dissociative electron attachment spectroscopy (DEAS). Three channels of molecular negative ions decay have been identified: abstraction of Cl⁻ and Cl₂⁻ as well as electron detachment (τ_a∼250 μs at 343 K). All three channels exhibit temperature dependence. The adiabatic electron affinity estimated using a simple but typically accurate Arrhenius model (EA_a=1.6–1.9 eV) turns out to be much higher than the quantum-chemical predictions (EA_a=0.9–1.0 eV). We discuss the possible reasons behind the observed discrepancy.     

Efficient biotinylation of methacryloyl-functionalized nonadherent cells for formation of cell microarrays

In order to obtain cell microarrays formed with human promyelocytic leukemia cells (HL-60), biotinylation of methacryloyl-functionalized HL-60 cells was performed via a thiol-ene reaction with thiol-terminated 4-arm poly(ethylene glycol) (PEG(4)10K-SH). Biotinylated HL-60 cells were selectively adhered onto an avidin-patterned surface with high viability.     

Attachment of metal trications to peptides

Gas-phase complexes of triply charged metal ions with peptides may be readily produced using electrospray ionization, including for small peptides such as bradykinin and peptides with no basic residues such as insulin chain A. Attachment without charge-reduction is demonstrated for all trications studied: La(3+), Al(3+), Ga(3+), Fe(3+), V(3+), and Cr(3+). The intensities of adducts are often comparable to, or even exceed, those of protonated analogs in any charge state.     

Attachment of drugs to polyethylene glycols

Polyethylene glycol (PEG) was used as a carrier polymer for the attachment, via end groups, of drugs such as penicillin V, aspirin, amphetamine, quinidine and atropine. For this purpose, methods were developed for the functionalization of PEG; PEG-NH₂, PEG-COOH and PEG-NCO were prepared. Use was made of dicyclohexyl carbodiimide together with 4-dimethylamino pyridine or 1-hydroxybenzotriazole for the coupling reactions.     

Assembly of multicellular constructs and microarrays of cells using magnetic nanowires

An approach is described for controlling the spatial organization of mammalian cells using ferromagnetic nanowires in conjunction with patterned micromagnet arrays. The nanowires are fabricated by electrodeposition in nanoporous templates, which allows for precise control of their size and magnetic properties. The high aspect ratio and large remanent magnetization of the nanowires enable suspensions of cells bound to Ni nanowires to be controlled with low magnetic fields. This was used to produce one- and two-dimensional field-tuned patterning of suspended 3T3 mouse fibroblasts. Self-assembled one-dimensional chains of cells were obtained through manipulation of the wires' dipolar interactions. Ordered patterns of individual cells in two dimensions were formed through trapping onto magnetic microarrays of ellipsoidal permalloy micromagnets. Cell chains were formed on the arrays by varying the spacing between the micromagnets or the strength of fluid flow over the arrays. The positioning of cells on the array was further controlled by varying the direction of an external magnetic field. These results demonstrate the possibility of using magnetic nanowires to organize cells.     

Electrophoretic deposition of bacterial cells

The possibility to use alternating current electrophoretic deposition (AC-EPD) to deposit living cells in the form of Staphylococcus aureus and Escherichia coli on stainless steel was assessed. The experimental results revealed that these bacteria can be successfully deposited on metallic surfaces from demineralized water and sucrose based solutions using asymmetric unbalanced electric fields. Cell viability of the deposited bacteria was influenced by the strain and deposition medium.     

Attachment of Gold Nanoparticles to Carbon Nanotubes

Carbon nanotubes were initially chemically modified with an H2SO4-HNO3 treatment, and subsequently activated with Pd-Sn catalytic nuclei via a one-step activation approach. These activated nanotubes were used as precursors for obtaining gold nanoparticles-attached nanotubes via simple electroless plating. This approach provides an efficient method for attachment of metal nanostructures to carbon nanotubes. Such novel hybrid nanostructures are attractive for many applications.     

Strategies to improve the surface plasmon resonance-based immmunodetection of bacterial cells

We have made a comparison of (a) different surface chemistries of SPR sensor chips (such as carboxymethylated dextran and carboxymethylated C1) and (b) of different assay formats (direct, sandwich and subtractive immunoassay) in order to improve the sensitivity of the determination of the model bacteria Acidovorax avenae subsp. citrulli (Aac). The use of the carboxymethylated sensor chip C1 resulted in a better sensitivity than that of carboxymethylated dextran CM5 in all the assay formats. The direct assay format, in turn, exhibits the best sensitivity. Thus, the combination of a carboxymethylated sensor chip C1 with the direct assay format resulted in the highest sensitivity for Aac, with a limit of detection of 1.6?×?10⁶ CFU mL^-1. This SPR immunosensor was applied to the detection of Aac in watermelon leaf extracts spiked with the bacteria, and the lower LOD is 2.2?×?10⁷ CFU mL^?1.

Relevance of electrokinetic theory for "soft" particles to bacterial cells: implications for bacterial adhesion

Bacterial cells and other biological particles carry charged macromolecules on their surface that form a "soft" ion-permeable layer. In this paper, we test the applicability of an electrokinetic theory for soft particles to characterize the electrophoretic mobility (EPM) and adhesion kinetics of bacterial cells. The theory allows the calculation of two parameters--the electrophoretic softness and the fixed charged density--that define the characteristics of the polyelectrolyte layer at the soft particle surface. The theory also allows the calculation of an outer-surface potential that may better predict the electrostatic interaction of soft particles with solid surfaces. To verify its relevance for bacterial cells, the theory was applied to EPM measurements of two well-characterized Escherichia coli K12 mutants having lipopolysaccharide (LPS) layers of different lengths and molecular compositions. Results showed that the obtained softness and fixed charge density were not directly related to the known characteristics of the LPS of the selected strains. Interaction energy profiles calculated from Derjaguin-Landau-Verwey-Overbeek (DLVO) theory were used to interpret bacterial deposition (adhesion) rates on a pure quartz surface. The outer surface potential failed to predict the low attachment efficiencies of the two bacterial strains. The lack of success in the application of the theory for soft particles to bacterial cells is attributed to chemical and physical heterogeneities of the polyelectrolyte layer at the cell surface.     

Sparse regularized discriminant analysis with application to microarrays

For cancer prediction using large-scale gene expression data, it often helps to incorporate gene interactions in the model. However it is not straightforward to simultaneously select important genes while modeling gene interactions. Some heuristic approaches have been proposed in the literature. In this paper, we study a unified modeling approach based on the ℓ₁ penalized likelihood estimation that can simultaneously select important genes and model gene interactions. We will illustrate its competitive performance through simulation studies and applications to public microarray data.     

Attachment of glycosaminoglycan oligosaccharides to thiol-derivatised gold surfaces

Glycosaminoglycan oligosaccharides have been attached to thiol-derivatised gold surfaces, via the formation of mercury-sugar adducts at the non-reducing end, representing a new method of generating versatile glycoconjugates incorporating this class of biologically and medically important carbohydrate.     

Application of click chemistry to the production of DNA microarrays

The copper-catalyzed alkyne-azide cycloaddition (CuAAC) reaction was applied as the novel method of DNA immobilization on a modified solid support. The CuAAC click reaction enables the covalent binding of DNA modified with pentynyl groups at its 5'-end to azide-loaded slides. Click microarrays were produced using this approach and successfully employed in biological/model experiments.