Nanofiber Grid and Related Methods

Patent metadata

Field	Value
Publication number	US-2016202289-A1
Application number	US-201614990206-A
Country	US
Kind code	A1
Filing date	Jan 7, 2016
Priority date	Jan 7, 2015
Publication date	Jul 14, 2016
Grant date	—

Abstract

Methods and systems are provided for measuring single and multi-cell inside-out and/or outside-in forces on a nanofiber grid. Single and multi-cells are deposited on, or migrate onto the nanofiber grid where the cell or cells are in contact with at least one fiber of the nanofiber grid and forces generated by the cells are observed and measured using deflection sensing methods. Furthermore, analyte-testing platforms using the nanofiber grid are described herein. Also provided are methods and apparatus including automated analyte-testing platforms using the nanofiber grid.

First claim

We claim:

1 . A method of measuring a cell force comprising:
a. providing one or more cells on a nanofiber grid suspended in an aqueous medium or a hydrogel, wherein the nanofiber grid comprises a plurality of high aspect ratio fibers having diameters of between about 10 nm and 10 μm, wherein the fibers are formed into a crossed pattern having one or more intersections, and wherein the fibers are fused at the intersections of the crossed pattern, wherein at least one cell is in contact with a first fiber; b. measuring deflection of the first fiber in contact with the at least one cell; and c. calculating from the deflection of the first fiber a force applied to the fiber by the at least one cell.

2 . The method of claim 1 , wherein the cell contacts a plurality of fibers and the deflection of more than one fiber is measured, and forces acting on the more than one fiber for which deflection is measured are calculated.

3 . The method of claim 1 , in which the high aspect ratio fibers are polymeric.

4 . The method of claim 3 , wherein the polymer is or more of a polystyrene, a polyester, a polyurethane, a polyacrylamide, a poly (methyl methacrylate), a polylactic acid, a poly(glycolic acid), a poly(lactic-co-glycolic acid), a polyaniline, a polypyrrole, fibrinogen, collagen, and mixtures and copolymers thereof, and/or includes carbon nanotubes, carbon black, or metallic nanoparticles.

5 . The method of claim 3 , in which the polymeric high aspect ratio fibers are prepared by determining an entanglement concentration (Ce) for a first polymer solution comprising a first polymer and a first good solvent for the first polymer; feeding the first polymer solution comprising the first polymer having a concentration of at least Ce in the first good solvent for the first polymer through a spinneret to produce an extruded droplet of polymer solution at a tip of the spinneret; contacting the extruded droplet of polymer solution with a target at a contact point; moving the contact point away from the spinneret, thereby pulling a high aspect ratio polymeric fiber from the extruded droplet of polymer solution at the tip of the spinneret; and further pulling the fiber from the extruded droplet of polymer solution at the tip of the spinneret and feeding the first polymer solution through the spinneret into the extruded droplet of polymer solution at the tip of the spinneret at a rate sufficient to compensate for an amount of the first polymer solution used to produce the fiber, thereby producing a bead-free, high aspect ratio polymeric fiber.

6 . The method of claim 1 , in which the nanofiber grid comprises a plurality of spaced-apart support fibers having a diameter ranging from 1 μm to 100 μm, spanning a frame, and a plurality of crossing fibers, crossing the support fibers, having a diameter of from 50 nm to 1 μm, and spaced-apart at a distance of between 10 μm and 100 μm.

7 . The method of claim 6 , in which the nanofiber grid comprises a frame, wherein the support fibers and the crossing fibers span the frame.

8 . The method of claim 6 , in which the crossing fibers form an angle with the support fibers of from 10° to 90°.

9 . The method of claim 6 , in which the support fibers are perpendicular to the crossing fibers.

10 . The method of claim 1 , wherein the high aspect ratio fibers formed into a crossed pattern having two different directions, wherein the high aspect ratio polymeric fibers have different diameters in each direction of the crossed pattern.

11 . The method of claim 1 , in which the first fiber deflects at least 20 nm with an applied force ranging from 10 pico-Newtons to 100 micro-Newtons, applied normal to the first fiber.

12 . The method of claim 1 , wherein the cell is attached to a second fiber, and the method further comprises, prior to measuring the deflection of the at least one fiber, moving a second fiber attached to the cell using a first probe placed at a point on the second fiber adjacent to the cell on a first side of the cell between the cell and a first intersection adjacent to the cell.

13 . The method of claim 12 , further comprising moving the second fiber using the first probe, and a second probe at a point on the second fiber adjacent to the cell on a second side of the cell opposite the first side between the cell and a second intersection adjacent to the cell.

14 . The method of claim 13 , wherein the force applied to the cell on the first side is different to the force applied to the cell on the second side.

15 . The method of claim 12 , further comprising moving the first fiber with a probe placed at a point on the first fiber adjacent to the cell on a side of the cell opposite a point on the first fiber at which the deflection of the fiber is measured between the cell and a second intersection adjacent to the cell.

16 . The method of claim 12 , in which deflection of the first fiber is indicative of cell-cell junction strength, cytoskeletal structure, cell integrity, cell stress and/or strain values, and/or cell drug response of the cell on the first fiber.

17 . The method of claim 12 , wherein the second fiber is moved until the cell begins to detach or detaches from the first fiber and/or the second fiber, and determining the force applied to the first fiber by the cell at the time the cell begins to detach and/or detaches from the first fiber and/or the second fiber.

18 . The method of claim 1 , in which one or more fibers of the nanofiber grid comprise a cell adhesion-promoting composition.

19 . The method of claim 18 , wherein the cell adhesion-promoting composition is one or more of: collagen, vitronectin, laminin, fibronectin, fibrinogen, poly(ornithine), poly(lysine), and a cell-adhesion promoting peptide.

20 . The method of claim 1 , in which the first fiber comprises a label.

21 . The method of claim 20 , in which the label is selected from the group consisting of a fluorescent dye, and a quantum dot.

22 . The method of claim 1 , in which deflection of the first fiber is measured by obtaining an image of the first fiber using a digital imaging device, transmitting the image of the first fiber to a computer, determining by use of a computer-implemented process the displacement of the first fiber by the cell, calculating from the displacement a force that is used to displace the first fiber to the extent depicted in the image, and producing an output indicating the force that is used to displace the first fiber to the extent depicted in the image.

23 . The method of claim 1 , further comprising adding one or more analytes to the aqueous medium or hydrogel and determining deflection of the first fiber either at one or more time points prior to or after addition of the active agent to the aqueous medium, or compared to a cell deposited on a second nanofiber grid in aqueous medium in a second vessel without addition of the active agent, or without addition of the same amount of active agent or sample.

24 . The method of claim 1 , further comprising aspirating the cell, wherein aspirating the cell comprises pulling the cell on the first fiber, and optionally detaching the cell from the first fiber, and wherein the deflection of the first fiber is measured during aspiration of the cell.

25 . The method of claim 1 , in which deflection of the first fiber and one or more additional fibers is measured to identify contraction forces and expansion forces of th

Assignees

• Univ Carnegie Mellon

Inventors

• Nain Amrinder Singh
• Behkam Bahareh

Classifications

• G01N33/4833 Primary

  of solid biological material, e.g. tissue samples, cell cultures (tissue in vivo A61B5/00; cell suspensions G01N33/48735) · CPC title

• G01Q30/04 Primary

  Display or data processing devices · CPC title