Micropatterning assembly, methods for micropatterning, and micropatterned devices

1 . A micropatterning system comprising a cell binding device for modular micropatterning one or more cell binding molecules comprising:

a) a solid carrier comprising a surface, wherein the surface is coated with a passivated polymeric coating that is covalently attached to the surface of the solid carrier; and

b) one or more adaptor molecules present in a liquid phase in contact with the passivated polymeric surface coating, each adaptor molecule comprising:

(i) a terminal photoreactive moiety configured to covalently photoimmobilize the adaptor molecule to a predetermined micropatterning area of the passivated polymeric coating of the carrier by photoinduced radical formation, wherein the terminal photoreactive moiety is a fluorescent dye, benzophenone, or phenylazide and

(ii) a terminal click chemistry cycloaddition moiety that is chemically distinct from and orthogonal to the photoreactive moiety, wherein the terminal click chemistry cycloaddition moiety is an alkyne, cyclic alkyne, transcycloocetene, norbornene, cyclopropane, or tetrazine, and the terminal click chemistry cycloaddition moiety is configured to covalently bind a counterpart reactive group on the one or more cell binding molecules by cycloaddition subsequent to photoimmobilization of the adaptor molecule by the terminal photoreactive moiety,

wherein the photoreactive moiety and the click chemistry cycloaddition moiety are covalently attached and spatially separated.

2 . The system of claim 1 , wherein the passivated polymeric coating comprises a hydrophilic synthetic polymer selected from the group consisting of polyvinyl alcohol (PVA), polyethylene glycol (PEG) and polyhydroxyethylmethacrylate (polyHEMA), and derivatives thereof.

3 . The system of claim 1 , wherein the fluorescent dye is fluorescein.

4 . The system of claim 1 , wherein the cell binding molecule further comprises a cell binding moiety recognized by a cellular surface structure or cell surface receptor, wherein the cell surface receptor is a cell adhesion receptor or a cell signaling receptor.

5 . The system of claim 4 , wherein the cell binding molecule is an extracellular signaling molecule.

6 . The system of claim 1 , wherein the cell binding molecules are covalently bound to the adaptor molecules in a continuous concentration gradient pattern on the solid carrier, wherein the gradient pattern is generated by sequential application of spatially resolved light exposure to different regions of the adaptor molecule-coated carrier surface.

7 . The system of claim 1 , which is any of an analytical, diagnostic, medical, or industrial device.

8 . A method for producing a system comprising a cell binding device for modular micropatterning one or more cell binding molecules comprising:

a) providing a solid carrier comprising a surface;

b) passivating the surface of the solid carrier by covalently attaching a polymeric coating to the surface of the solid carrier;

c) contacting the passivated surface of the solid carrier with a liquid phase comprising one or more adaptor molecules, wherein each of the one or more adaptor molecules comprises:

(i) a terminal photoreactive moiety configured to covalently photoimmobilize the adaptor molecule to a predetermined micropatterning area of the passivated polymeric coating on the solid carrier surface by photoinduced radical formation, wherein the terminal photoreactive moiety is a fluorescent dye, benzophenone, or phenylazide, and

(ii) a terminal click chemistry cycloaddition moiety that is chemically distinct from and orthogonal to the photoreactive moiety, wherein the terminal click chemistry cycloaddition moiety is an alkyne, cyclic alkyne, transcycloocetene, norbornene, cyclopropane, or tetrazine, and the terminal click chemistry cycloaddition moiety is configured to covalently bind a counterpart reactive group present on the one or more cell binding molecules by cycloaddition subsequent to photoimmobilization of the adaptor molecule by the terminal photoreactive moiety,

wherein the photoreactive moiety and the click chemistry cycloaddition moiety are covalently attached and spatially separated;

d) projecting a desired light pattern onto the solid carrier surface at the liquid interface to selectively activate the photoreactive moiety, thereby covalently immobilizing the adaptor molecules to predetermined micropatterning areas of the passivated polymeric coating while maintaining the click chemistry cycloaddition moiety in a chemically unaltered and available state for subsequent covalent binding; and

e) contacting the one or more immobilized adaptor molecules with the one or more cell binding molecules under conditions that enable the click chemistry cycloaddition moiety to covalently bind with the counterpart reactive group on the cell binding molecules by cycloaddition, thereby producing a micropatterned cell binding device.

9 . The system of claim 1 , wherein a preparation of bioactive target cells is specifically bound to the device.

10 . The system of claim 1 , wherein the target cells are selected from epithelial cells, tumor cells, leukocytes, mesenchymal cells, and stem cells.

11 . The system of claim 1 , wherein the counterpart reactive group present on the cell binding molecule is selected from the group consisting of: a terminal azide, a terminal alkyne, cyclic alkyne, trans-cyclooctene, norbornene, cyclopropane, and tetrazine.

12 . The system of claim 4 ,

wherein the adhesion receptors are integrins, cadherins, selectins or immunoglobulins, and wherein the cell signaling receptors are G-protein coupled receptors, receptor tyrosine kinases, receptor serine/threonine kinases, receptor guanylyl cyclases, and histidine kinase associated receptors.

13 . The system of claim 5 , wherein the extracellular signaling molecule is a peptide comprising any of the RGD motif derivatives, formyl-methionyl-leucyl-phenylalanine (fMLP), chemokines, G-protein coupled receptor ligands, receptor tyrosine kinase ligands, receptor serine/threonine kinase ligands, receptor guanylyl cyclase ligands, and histidine kinase 64 receptor ligands.

14 . The system of claim 7 , wherein the device is selected from a microscopy slide, affinity matrix, cell culture support, diagnostic array, medical implant, and cell migration applications.

15 . The system of claim 14 , wherein the cell migration applications are selected from chemotaxis applications, haptotaxis applications, and microfluidic applications.

16 . The system of claim 9 , wherein the target cells are specifically binding as a monolayer and/or cell clusters.