Nanoelectromechanical system (NEMS) force sensors, when applied to biological applications, can measure the force exerted by a single cell on its surroundings with unparalleled sensitivity. A NEMS force sensor and microfluidic device for screening one or more cells have been developed by California Institute of Technology(Pasadena, CA) scientists.
According to U. S. Patent Application 20100041091, the microfluidic device includes: (i) providing one or more cells to a nanoelectromechanical system (NEMS) force sensor; (ii) applying at least one reagent to the one or more cells; and (iii) observing a response of the one or more cells to the reagent with the force sensor, thereby screening the one or more cells. The device and method were developed by Professor of Physics, Applied Physics, and Bioengineering at the Caltech and codirector of Caltech's Kavli Nanoscience Institute Michael L. Roukes, Condensed Matter Physics Research Engineer Blake W. Axelrod and Staff Scientist Jessica L. Arlett.
The scientists also developed a method of evaluating effects of a drug reagent, comprising: (i) providing one or more cells to a nanoelectromechanical system (NEMS) force sensor; (ii) applying the drug reagent to the one or more cells; and (iii) observing a response of the one or more cells to the drug reagent with the force sensor, whereby an effect of the drug reagent on the one or more cells is evaluated.
The scientists also developed a method of evaluating effects of a drug reagent, comprising: (i) providing one or more cells to a nanoelectromechanical system (NEMS) force sensor; (ii) applying the drug reagent to the one or more cells; and (iii) observing a response of the one or more cells to the drug reagent with the force sensor, whereby an effect of the drug reagent on the one or more cells is evaluated.
The NEMS devices can be fabricated from lithographically patternable polymer instead of traditional MEMS materials, such as silicon, because polymer's low Young's modulus can enable improved sensitivity and fabrication of devices that are compliance-matched to typical biological materials. Additionally, the material costs for polymers can be significantly less than traditional semiconductor materials. This can be significant, particularly when, for example, the force sensor system comprises an array of sensors.
One advantage of NEMS force sensors for phenotypic screening can be the high sensitivity to perturbations of the cellular cytoskeleton. NEMS force sensors can be sensitive to subtle effects of potential reagents that other screening methods will miss--this can allow for detection of subtle activity that is desired in potential pharmaceuticals.
Additionally, NEMS force sensors can be compatible with microfluidic systems, thereby reducing the quantity of reagent, and thus cost, needed for a screening test. Microfluidic compatibility also can enable a degree of system automation and parallelization desirable for high throughput screening. Large arrays of NEMS force sensors can be fabricated and integrated for high throughput screening. Moreover, NEMS force sensors can be integrated with fluorescent microscopy. This enables molecular level monitoring of activity within a cell, a second method of monitoring cellular physiology and a very common tool used in high-content pharmaceutical screening.
In one embodiment, a long-trench-shaped cell chamber, which is spanned by the force sensing beams (or "force sensor") and a bridge to hold the cell being studied in close proximity to the beams can be made. The chamber can have any suitable size. For example, in one embodiment it can be roughly 600-800 microns long and 110 microns wide, or it can be longer or shorter, wider or narrower, depending on the use.
The cantilevers (or "beams") of the force sensor can be fabricated from two layers of polymer that can sandwich a strain sensor (or "strain gauge"), thus keeping the strain sensor from the surrounding liquid and symmetric--decoupled--with respect to out-of-plane deflections. The liquid can come from a microfluidics channel that can be embedded and integrated with the system. The microfluidics can, for example, encapsulate the force sensor. The polymer layer can be, for example, between 50 nm and 200 nm thick, such as about 80 nm to about 150 nm thick, such as about 100 nm thick.
FIGS. 1a-1b show images and schematic illustrating how a cell deforms a force sensing beam upon contraction. FIG. 1a provides electron micrographs of suspended beams (FIG. 1a(i)), a zoom-in image thereof (FIG. 1a(ii)), and an optical image of microfluidics encapsulated chip held between thumb and forefinger (FIG. 1a(iii)). FIG. 1B provides schematics showing a cell attaching and spreading on the large center gold pad then reaching the force sensors on both sides (FIG. 1b(i)), causing lengthening of the wires.
FIGS. 1a-1b show images and schematic illustrating how a cell deforms a force sensing beam upon contraction. FIG. 1a provides electron micrographs of suspended beams (FIG. 1a(i)), a zoom-in image thereof (FIG. 1a(ii)), and an optical image of microfluidics encapsulated chip held between thumb and forefinger (FIG. 1a(iii)). FIG. 1B provides schematics showing a cell attaching and spreading on the large center gold pad then reaching the force sensors on both sides (FIG. 1b(i)), causing lengthening of the wires.
FIGS. 2a-2i illustrate NEMS-enabled single-cell force measurements with high temporal and force resolution. FIGS. 2a-2b provide optical images showing a microfluidics encapsulated force sensor, and FIG. 2c provides an electron micrograph thereof. FIG. 2d shows an NIH-3T3 cell attached to a microfluidics encapsulated force sensor in a microscope-mounted incubator and sample holder. FIG. 2e shows force versus time data from a contracting and relaxing lamellipodium, as the cell is perturbed with cytochalasin D and allowed to recover in growth media (as shown in FIG. 2b).
The force data was acquired with a force resolution of 200 pN and a time resolution of 100 ms--25.times. and 300.times., respectively, better than prior state of the art. FIGS. 2f-2h provide plots that show force signatures of two molecular-mechanical processes: small force oscillations, roughly 400 pN peak-to-peak, with a frequency that is monotonically dependent on the force being exerted by the lamellipodium shown in FIG. 2f, with FIG. 2g illustrating the linearity of frequency dependence.
FIGS. 2h-2i show that large, stable, quantized force steps of order 1 nN are manifested when a cell's cytoskeleton is perturbed with cytochalasin D and allowed to recover in growth media. A histogram of the step sizes is presented in FIG. 2j.
FIGS. 3a-3d show results from surface chemistry Petri dish experiments. FIG. 3a shows results of a typical fibronectin only test: cells are spread everywhere. FIG. 3b is from a Pluronics only test: almost no cells present and none are spread. FIG. 3c-3d show results from complete surface chemistry process (SAM, Pluronics and fibronectin): cells are well spread on the gold and absent from the polymer.
A microfluidics channel can be integrated with the force sensor system. For example, it can encapsulates the force sensor, wherein the microfluidics channel can control the fluidic environment around the force sensor. One advantage of such design is that the microfluidics can improve the electrical read out of the electrical signal from the sensor
Polymer NEMS-based force sensors that directly measure the force a cell under study exerts on its surroundings with near-single molecule force resolution and whole cell dynamic range have been developed by Caltech’s Roukes Group
A microfluidics embedded NEMS force sensor can be fabricated by first depositing a polymer layer onto a silicon nitride membrane layer. The membrane layer can be disposed over a silicon wafer. A metal wire, or a strain sensor, comprising a piezoresistive element can be deposited onto the polymer layer. An additional polymer layer can then be deposited onto the metal wire. The polymer layers need not comprise the same material, but can, if suitable.
The polymer can comprise any polymer, including parylene, SU-8, polyimide, or combinations thereof. The backside of the wafer can be etched to create a hole. As a result of this process, in this embodiment, a NEMS force sensor can be fabricated from two layers of polymer, encapsulating a metal wire. The force sensor is suspended over a hole, with a portion of the silicon-nitride membrane removed without damage to the polymer layers.
Polymer NEMS-based force sensors that directly measure the force a cell under study exerts on its surroundings with near-single molecule force resolution and whole cell dynamic range have been developed by Caltech’s Roukes Group
Last year the group of Caltech physicists created first nanoscale mass spectrometer. The new technique, developed over 10 years of effort by Michael L. Roukes, Professor of Physics, Applied Physics, and Bioengineering at the Caltech and codirector of Caltech's Kavli Nanoscience Institute, and his colleagues, simplifies and miniaturizes the process through the use of very tiny nanoelectromechanical system (NEMS) resonators.
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