Biomedical subjects

Stephen C Jacobson

Publications and source records attributed to Stephen C Jacobson.

10 recordsLinked to original sources

PubMed2006

Fabrication of three-dimensional micro- and nanoscale features with single-exposure photolithography.

We report a technique for fabricating three-dimensional structures from two-dimensional photomasks in a single exposure. Size-dependent transmission properties of apertures in the photomask and exposure energy were used to control polymer feature dimensions. The photomasks were produced by electron beam lithography, and apertures in the photomasks were 0.35-5.5 microm wide and 20-30 microm long. Photomasks were coated with the negative tone photoresist SU-8, and following exposure and postexposure processing, the resulting SU-8 features had widths from 0.35 to 5.5 microm and heights from 1.1 to 10.8 microm. With this technique, nanoscale features were easily coupled to microscale features because they were created in the same photoresist layer with one exposure.

Microscopy, Electron, Scanning↗

PubMed2006

Chemotaxis assays of mouse sperm on microfluidic devices.

Sperm chemotaxis is an area of significant interest to scientists involved in reproductive science. Understanding how and when sperm cells are attracted to the egg could have profound effects on reproduction and contraception. In an effort to systematically study this problem, we have fabricated and evaluated a microfluidic device to measure sperm chemotaxis. The device was designed with a flow-through configuration using a spatially and temporally stable chemical gradient. Mouse sperm cells were introduced into the chemotaxis chamber between confluent flows of mouse ovary extract and buffer. The sperm experiencing chemotaxis swam toward the extract and were counted relative to those that swam toward the buffer. The ovary extracts were diluted from 10(2) to 10(7) times, and each extract dilution was screened for chemotaxis. Four out of six ovaries showed a strong chemotactic response at extract dilutions of 10(-3) to 10(-5). This device provided a convenient, disposable platform on which to conduct chemotaxis assays, and the flow-through design overcomes difficulties associated with distinguishing chemotaxis from trapping.

Animals↗

PubMed2005

Stacking due to ionic transport number mismatch during sample sweeping on microchips.

Sample stacking can occur in isoconductive buffer systems as a result of ion transport mismatches that cause changes in buffer conductivity during electrophoresis. Fluorescence imaging was used to examine this effect in the sweeping of hydrophobic dyes with sodium dodecyl sulfate (SDS) on microchips. Imaging revealed the occurrence of a stacking effect in a sodium borate buffer system in which the sample buffer and SDS-containing run buffer had the same initial conductivity. Injected sample plugs were first swept by SDS micelles and the swept band was then stacked at the trailing end of the sample zone. This effect is due to changes in conductivity at both the front and back interfaces of the injected sample plug and can be modeled by moving boundary equations. Maximum signal enhancements of 86-, 160- and 560-fold were obtained for Rhodamine 560, Rhodamine B and Rhodamine 6G, respectively, by the combination of sweeping and stacking within a 1 cm section of microchannel. Based on sample sweeping/stacking and manipulation of the electric field polarity, a method of trapping and concentrating analyte from multiple injections was also demonstrated.

Buffers↗

PubMed2005

Static and dynamic acute cytotoxicity assays on microfluidic devices.

Static and dynamic acute toxicity assays of cells were performed on microfluidic devices where materials were hydraulically transported. Static assays were performed by incubating cells with an agent in a microchip reservoir and optically interrogating the cells after hydrodynamic focusing at a cross intersection. Dynamic assays were performed on a microchip with a 25-cm-long spiral channel where the cells were mixed with an agent and optically monitored 0.1, 12, and 22 cm from the point of mixing. The incubation time was determined by the time needed for cells to transit from the mixing location to the point of detection. Cell viability was determined using the ratio of fluorescence signals from membrane permeant (calcein) and membrane impermeant (propidium iodide) stains. The model system used in this study was the viability of Jurkat cells in the presence of the agent Triton X-100). An average LC50 value of 138 microM for Triton X-100 was obtained for an incubation period of 7-12 min using the static assay. LC50 values obtained with the dynamic assay for 25- and 47-s incubation times were 290 and 250 microM Triton X-100, respectively. Higher LC50 values for the dynamic assay were expected due to the shorter incubation times.

Cell Survival↗

PubMed2005

Preconcentration of proteins on microfluidic devices using porous silica membranes.

Fluorescently labeled proteins were electrophoretically concentrated on microfabricated devices prior to separation and laser-induced fluorescence detection on the same device. The proteins were concentrated using a porous silica membrane between adjacent microchannels that allowed the passage of buffer ions but excluded larger migrating molecules. Concentrated analytes were then injected into the separation column for analysis. Two basic microchip designs were tested that allowed sample concentration either directly in the sample injector loop or within the microchannel leading from the sample reservoir to the injector. Signal enhancements of approximately 600-fold were achieved by on-chip preconcentration followed by SDS-CGE separation. Preconcentration for CE analysis in both coated and uncoated open channels was also demonstrated. Fluorescently labeled ovalbumin could be detected at initial concentrations as low as 100 fM by using a combination of field-amplified injection and preconcentration at a membrane prior to CE in coated channels.

Electrophoresis, Capillary↗

PubMed2005

Three-dimensional mapping of the light intensity transmitted through nanoapertures.

A general method to map the 3D spatial distribution of light emerging from nanoscale apertures is presented that uses photolithographic techniques to create polymer replicas of the intensity distribution. The resulting features varied with aperture diameter and exposure time and showed good correlation with theory. This method provides direct visualization of the intensity distribution in close proximity to nanostructures and overcomes limitations imposed by physical probes where the contribution of the probe to the map requires deconvolution.

Computer Simulation↗

PubMed2004

Strategy for repetitive pinched injections on a microfluidic device.

A microfluidic valve was fabricated with a cross intersection and two tee intersections in close proximity and evaluated for repetitive pinched injections. Electrokinetic forces were used to mobilize the sample and control diffusive transport at a cross intersection to produce sample plugs of short axial extent in an analysis channel similar to the standard pinched valve. The addition of a tee intersection in the sample channel maintained the sample close to the injection valve under "pullback"conditions allowing more rapid loading into the cross intersection. A second tee intersection allowed unidirectional transport in the analysis channel enabling loading of subsequent injections during an analysis. The two tee intersections were each located 80 microm from the cross intersection. Injection frequencies of 1, 2.5, 5, and 10 Hz were tested with a duty cycle of 0.5 for sample loading and dispensing. With 1 kV applied to the microchip during dispensing, the relative standard deviation of the peak areas for 15 injections was 1.6%. The peak width (4sigma) for the repetitive injections increased from 71 to 96 microm compared to a standard pinched injection due to the presence of the tee intersection in the analysis channel.

Electrochemistry↗

PubMed2003

Microfluidic devices for the high-throughput chemical analysis of cells.

A microfluidic device is reported that integrated cell handling, rapid cell lysis, and electrophoretic separation and detection of fluorescent cytosolic dyes. The device function was demonstrated using Jurkat cells that were loaded with the fluorogenic dyes - carboxyfluorescein diacetate, Oregon green carboxylic acid diacetate, or Calcein AM. The loaded cells were hydrodynamically transported from the cell-containing reservoir to a region on the microfluidic device where they were focused and then rapidly lysed using an electric field. Complete lysis was accomplished in <33 ms. The hydrolyzed, fluorescent dyes in the cell lysate were automatically injected into a separation channel on the device and detected 3 mm downstream of the injection point. The total separation time was approximately 2.2 s with absolute migration time reproducibilities of <1% and efficiencies ranging from 2300 to 4000 theoretical plates. Results from 139 cells are reported. A small fraction of these cells, approximately 9%, were found to enzymatically hydrolyze the loaded dyes in a manner significantly different from the majority of the cells. Cell analysis rates of 7-12 cells/min were demonstrated and are >100 times faster than those reported using standard bench-scale capillary electrophoresis.

Algorithms↗

PubMed2003

High-efficiency, two-dimensional separations of protein digests on microfluidic devices.

High-efficiency, two-dimensional separations of tryptic digests were achieved using glass microfluidic devices. Following micellar electrokinetic chromatography (MEKC) separations in a 19.6-cm-long serpentine channel, the peptides were rapidly sampled into a 1.3-cm-long second-dimension channel, where they were separated by capillary electrophoresis (CE). The turns in the serpentine channel were asymmetrically tapered to minimize geometrical contributions to band broadening and to provide ample channel length for high-efficiency chromatographic separations. Analysis of rhodamine B injections routinely produced plate numbers of 230000 and 40000 in the first (MEKC) and second (CE) dimensions, respectively, corresponding to plate heights of 0.9 and 0.3 microm. The electric field strengths were 200 V/cm for MEKC and 2400 V/cm for CE. In analysis times less than 15 min, two-dimensional separation of bovine serum albumin tryptic digest produced a peak capacity of 4200 (110 in the first dimension and 38 in the second dimension). The system was used to identify a peptide from a tryptic digest of ovalbumin using standard addition and to distinguish between tryptic digests of human and bovine hemoglobin.

Animals↗

PubMed2003

Sample filtration, concentration, and separation integrated on microfluidic devices.

Microfabricated devices integrating sample filtration, solid-phase extraction, and chromatographic separation with solvent programming were demonstrated. Filtering of the sample was accomplished at the sample inlet with an array of seven channels each 1 microm deep and 18 microm wide. Sample concentration and separation were performed on channels 5 microm deep and 25 microm wide coated with a C18 phase, and elution was achieved under isocratic, step, or linear gradient conditions. For the solid-phase extraction, signal enhancement factors of 400 over a standard injection of 1.0 s were observed for a 320-s injection. Four polycyclic aromatic compounds were resolved by open channel electrochromatography in under 50 s. Chip operation was unaffected by the presence of the 5-microm silica particles at the filter entrance.

Journal Article↗