Johan G. Bomer

The micro-Petri dish, a million-well growth chip for the culture and high-throughput screening of microorganisms

A miniaturized, disposable microbial culture chip has been fabricated by microengineering a highly porous ceramic sheet with up to one million growth compartments. This versatile culture format, with discrete compartments as small as 7 × 7 μm, allowed the growth of segregated microbial samples at an unprecedented density. The chip has been used for four complementary applications in microbiology. (i) As a fast viable counting system that showed a dynamic range of over 10,000, a low degree of bias, and a high culturing efficiency. (ii) In high-throughput screening, with the recovery of 1 fluorescent microcolony in 10,000. (iii) In screening for an enzyme-based, nondominant phenotype by the targeted recovery of Escherichia coli transformed with the plasmid pUC18, based on expression of the lacZ reporter gene without antibiotic-resistance selection. The ease of rapid, successive changes in the environment of the organisms on the chip, needed for detection of β-galactosidase activity, highlights an advantageous feature that was also used to screen a metagenomic library for the same activity. (iv) In high-throughput screening of >200,000 isolates from Rhine water based on metabolism of a fluorogenic organophosphate compound, resulting in the recovery of 22 microcolonies with the desired phenotype. These isolates were predicted, on the basis of rRNA sequence, to include six new species. These four applications suggest that the potential for such simple, readily manufactured chips to impact microbial culture is extensive and may facilitate the full automation and multiplexing of microbial culturing, screening, counting, and selection.

Al2O3/Silicon NanoISFET with Near Ideal Nernstian Response

Nanoscale ISFET (ion sensitive field-effect transistor) pH sensors are presented that produce the well-known sub-nernstian pH-response for silicon dioxide (SiO(2)) surfaces and near ideal nernstian sensitivity for alumina (Al(2)O(3)) surfaces. Titration experiments of SiO(2) surfaces resulted in a varying pH sensitivity ∼20 mV/pH for pH near 2 and >45 mV/pH for pH > 5. Measured pH responses from titrations of thin (15 nm) atomic layer deposited (ALD) alumina (Al(2)O(3)) surfaces on the nanoISFETs resulted in near ideal nernstian pH sensitivity of 57.8 ± 1.2 mV/pH (pH range: 2-10; T = 22 °C) and temperature sensitivity of 0.19 mV/pH °C (22 °C ≤ T ≤ 40 °C). A comprehensive analytical model of the nanoISFET sensor, which is based on the combined Gouy-Chapman-Stern and Site-Binding (GCS-SB) model, accompanies the experimental results and an extracted ΔpK ≈ 1.5 from the measured responses further supports the near ideal nernstian pH sensitivity.

Chemically modified field-effect transistors; a sodium ion selective sensor based on calix[4]arene receptor molecules

The development of an ion-sensitive field-effect transistor for sodium ions is described. Cahx[4]arene derivatives incorporated in a poly(vinyl chloride)-based membrane provide the selectivity. A poly(2-hydroxyethyl methacrylate) interlayer between the silicon dioxide gate and the sensing membrane is necessary to obtain a Na+-sensitive ISFET with Nernstian behaviour. The potentiometric selectivity coefficients (log Kij pot) for Na+ over K+ and Li+ are ?1.9 and ?2.5,

Microfluidics with ultrasound-driven bubbles

Microstreaming from oscillating bubbles is known to induce vigorous vortex flow. Here we show how to harness the power of bubble streaming in an experiment to achieve directed transport flow of high velocity, allowing design and manufacture of microfluidic MEMS devices. By combining oscillating bubbles with solid protrusions positioned on a patterned substrate, solid beads and lipid vesicles are guided in desired directions without microchannels. Simultaneously, the flow exerts controlled localized forces capable of opening and reclosing lipid membranes.

Top-down fabrication of Sub-30 nm Monocrystalline Silicon nanowires using conventional microfabrication

We report a new low-cost top-down silicon nanowire fabrication technology requiring only conventional microfabrication processes including microlithography, oxidation, and wet anisotropic plane-dependent etching; high quality silicon nanowire arrays can be easily made in any conventional microfabrication facility without nanolithography or expensive equipment. Silicon nanowires with scalable lateral dimensions ranging from 200 nm down to 10-20 nm and lengths up to approximately 100 microm can be precisely formed with near-perfect monocrystalline cross sections, atomically smooth surfaces, and wafer-scale yields greater than 90% using a novel size reduction method where silicon nanowires can be controllably scaled to any dimension and doping concentration independent of large contacting regions from a continuous layer of crystalline silicon.

Electrokinetic label-free screening chip: a marriage of multiplexing and high throughput analysis using surface plasmon resonance imaging

We present an electrokinetic label-free biomolecular screening chip (Glass/PDMS) to screen up to 10 samples simultaneously using surface plasmon resonance imaging (iSPR). This approach reduces the duration of an experiment when compared to conventional experimental methods. This new device offers a high degree of parallelization not only for analyte samples, but also for multiplex analyte interactions where up to 90 ligands are immobilized on the sensing surface. The proof of concept has been demonstrated with well-known biomolecular interactant pairs. The new chip can be used for high throughput screening applications and kinetics parameter extraction, simultaneously, of interactant-protein complex formation.

Chemically modified field effect transistors: the effect of ion-pair association on the membrane potentials

A theoretical model has been developed which relates physically accessible parameters to the formation of a membrane potential. The description is an extension of a theoretical description presented previously by our group, now including divalent cations and ion-pair association. Simulations of the overall membrane potential reveal several factors that may lead to non-Nernstian response curves. For monovalent and divalent cations a reduction in the slope of the response curve (sub-Nernstian response) should virtually always be expected when ion-pair association takes place in the membrane. Ion-pair association of divalent cations and sample anions can lead to a super-Nernstian response. A diffusion potential generally reduces the Nernstian slope of the response curve. In addition, several experimental results are described which illustrate and confirm our theoretical model.

Planar interdigitated electrolyte-conductivity sensors on an insulating substrate covered with Ta2O5

Interdigitated electrolyte-conductivity sensors with an added top layer of insulating Ta2O5 have been realized. The electrode-substrate structure under the Ta2O5 film has been planarized in order to obtain a totally flat top surface. In addition, the electrodes have been applied on a totally insulating substrate, thus reducing the parasitic sensor capacitance by a factor of ten.

Parallel single-cell analysis microfluidic platform.

We report a PDMS microfluidic platform for parallel single‐cell analysis (PaSCAl) as a powerful tool to decipher the heterogeneity found in cell populations. Cells are trapped individually in dedicated pockets, and thereafter, a number of invasive or non‐invasive analysis schemes are performed. First, we report single‐cell trapping in a fast (2–5 min) and reproducible manner with a single‐cell capture yield of 85% using two cell lines (P3x63Ag8 and MCF‐7), employing a protocol which is scalable and easily amenable to automation. Following this, a mixed population of P3x63Ag8 and MCF‐7cells is stained in situ using the nucleic acid probe (Hoechst) and a phycoerythrin‐labeled monoclonal antibody directed at EpCAM present on the surface of the breast cancer cells MCF‐7 and absent on the myeloma cells P3x63Ag8 to illustrate the potential of the device to analyze cell population heterogeneity. Next, cells are porated in situ using chemicals in a reversible (digitonin) or irreversible way (lithium dodecyl sulfate). This is visualized by the transportation of fluorescent dyes through the membrane (propidium iodide and calcein). Finally, an electrical protocol is developed for combined cell permeabilization and electroosmotic flow (EOF)‐based extraction of the cell content. It is validated here using calcein‐loaded cells and visualized through the progressive recovery of calcein in the side channels, indicating successful retrieval of individual cell content.

Rapid sacrificial layer etching for the fabrication of nanochannels with integrated metal electrodes

We present a rapid etch method to surface-micromachine nanochannels with integrated noble metal electrodes using a single metal sacrificial layer. The method is based on the galvanic coupling of a chromium sacrificial layer with gold electrodes, which results in a 10-fold increase in etch rate with respect to conventional single metal etching. The etch process is investigated and characterized by optical and electrochemical measurements, leading to a theoretical explanation of the observed etch rate based on mass transport. Using this explanation we derive some generic design rules for nanochannel fabrication employing sacrificial metal etching.