Capture, purification, and release of biological substances using a surface coating

Abstract

This invention relates to a surface coating for capture circulating rare cells, comprising a nonfouling composition to prevent the binding of non-specific cells and adsorption of serum components; a bioactive composition for binding the biological substance, such as circulating tumor cells; with or without a linker composition that binds the nonfouling and bioactive compositions. The invention also provide a surface coating for capture and purification of a biological substance, comprising a releasable composition to release the non-specific cells and other serum components; a bioactive composition for binding the biological substance, such as circulating tumor cells; with or without a linker composition that binds the releasable and bioactive compositions. The present invention also discloses a novel microfluidic chip, with specific patterned microstructures to create a flow disturbance and increase the capture rate of the biological substance.

What is claimed is: 
     
         1 - 63 . (canceled) 
     
     
         64 . A microfluidic chip for selectively enriching rare cells, the microfluidic chip comprising a substrate and a surface coating for capturing rare cells, wherein the surface coating comprises:
 (i) a bioactive composition which selectively binds to the rare cells, wherein the bioactive composition comprises an antibody; and   (ii) a non-fouling composition that reduces binding of non-specific cells or adsorption of serum proteins compared to a surface coating lacking the non-fouling composition, wherein the non-fouling composition comprises a lipid layer; and   wherein the antibody is non-covalently associated with the non-fouling composition.   
     
     
         65 . The microfluidic chip of  claim 64 , wherein the surface coating is non-covalently associated with the substrate. 
     
     
         66 . The microfluidic chip of  claim 65 , wherein the non-fouling composition is disposed between the bioactive composition and the substrate. 
     
     
         67 . The microfluidic chip of  claim 65 , wherein the non-fouling composition is coupled to the substrate by a linker. 
     
     
         68 . The microfluidic chip of  claim 64 , wherein the antibody is non-covalently associated with the non-fouling composition via a non-covalent interaction selected from the group consisting of hydrogen bonding, electrostatic interaction, hydrophilic-hydrophilic interaction, polar-polar interaction, magnetic force, and combinations thereof. 
     
     
         69 . The microfluidic chip of  claim 64 , wherein the lipid layer is a lipid bilayer. 
     
     
         70 . The microfluidic chip of  claim 69 , wherein the lipid bilayer is a supported lipid bilayer. 
     
     
         71 . The microfluidic chip of  claim 64 , wherein the substrate comprises a series of microstructures configured to interact with cells, wherein the series of microstructures are perpendicular to a flow direction of the microfluidic chip. 
     
     
         72 . The microfluidic chip of  claim 71 , wherein the series of microstructures comprises microstructures arranged in a linear pattern such that microstructures in a first pair of adjacent rows have a distance separating the adjacent rows, thereby forming a first gap, and wherein said first gap is not in line with a second gap formed by a second pair of adjacent rows, wherein said first gap and second gap are in adjacent columns. 
     
     
         73 . The microfluidic chip of  claim 64 , wherein the antibody is a biotinylated antibody. 
     
     
         74 . The microfluidic chip of  claim 64 , wherein the antibody is an EpCAM antibody. 
     
     
         75 . The microfluidic chip of  claim 74 , wherein the EpCAM antibody comprises a heavy chain and a light chain, wherein
 (a) the heavy chain comprises CDR1, CDR2, and CDR3 of SEQ ID NO: 1, and   (b) the light chain comprises CDR1, CDR2, and CDR3 of SEQ ID NO: 2.   
     
     
         76 . The microfluidic chip of  claim 64 , further comprising a syringe pump configured to apply buffer at a flow rate configured to release non-specific cells from the non-fouling layer without releasing cells selectively bound to the bioactive composition. 
     
     
         77 . The microfluidic chip of  claim 64 , further comprising a syringe pump configured to aid rinsing the microfluidic chip with a buffer at a shear force of about 2.5 to about 10 dyne/cm 2 . 
     
     
         78 . The microfluidic chip of  claim 64 , wherein the bioactive composition or the surface coating is configured to be removed from the substrate by flow of a buffer comprising bubbles. 
     
     
         79 . The microfluidic chip of  claim 64 , wherein the surface coating is configured such that flow of a buffer at shear force of less than about 50 dyne/cm 2  releases non-specificcells from the non-fouling layer without releasing cells selectively bound to the bioactive composition. 
     
     
         80 . The microfluidic chip of  claim 79 , wherein the surface coating is configured such that flow of a buffer at a shear force of from about 2.5 dyne/cm 2  to about 10 dyne/cm 2  releases non-specific cells from the non-fouling layer without releasing cells selectively bound to the bioactive composition. 
     
     
         81 . The microfluidic chip of  claim 64 , wherein the surface coating is configured such that flow of a buffer at a shear force of 50 dyne/cm 2  or more releases the bioactive composition or the surface coating from the substrate. 
     
     
         82 . A method of making a microfluidic chip for use in selectively enriching rare cells, the method comprising applying a bioactive composition which selectively binds to the rare cells to a non-fouling layer associated with a substrate of the microfluidic chip, wherein the non-fouling layer comprises a lipid layer and the bioactive composition comprises an antibody, wherein the antibody non-covalently associates with the non-fouling layer. 
     
     
         83 . A method of collecting rare cells from a biological sample, the method comprising:
 (a) contacting the biological sample to a microfluidic chip comprising a substrate and a surface coating for the capturing rare cells, wherein the surface coating comprises:
 (i) a bioactive composition which selectively binds to the rare cells, wherein the bioactive composition comprises an antibody; and 
 (ii) a non-fouling composition that reduces binding of non-specific cells or adsorption of serum proteins compared to a surface coating lacking the non-fouling composition; 
   (b) binding the rare cells to the antibody;   (c) flowing a buffer at shear stress of from about 2.5 dyne/cm 2  to about 12 dyne/cm 2  through the microfluidic chip, thereby releasing non-specific cells from the non-fouling layer without releasing the rare cells bound to antibody from the microfluidic chip, wherein the antibody remains non-covalently associated with the non-fouling lipid layer; and   (d) flowing a buffer at shear stress at least 50 dyne/cm 2  through the microfluidic chip, thereby releasing the rare cells and the bioactive composition or the surface coating from the substrate.