Functionalized 3D-Hydrogel Plugs Covalently Patterned Inside Hydrophilic Poly(dimethylsiloxane) Microchannels for Flow-Through Immunoassays

Abstract

Integration of a hydrogel and polydimethylsiloxane (PDMS)-based microfluidic device can greatly reduce the cost of developing channel-based devices. However, there are technical difficulties including the hydrophobic and inert surface properties associated with PDMS as well as back pressure and fragile material associated with the use of hydrogel in microchannels. In this study, a strategy to covalently photopattern 3-D hydrogel plugs with functionalized protein G inside microfluidic channels on a hydrophilic PDMS substrate coated with polyelectrolyte multilayers (PEMS) is presented. In this process, a UV-light microscope is applied to initiate the protein G-poly(acryl amide) copolymerization from the bulk substrate to solution areas via the deeply implanted photoinitiator (PI), resulting in sturdy 3D plugs covalently bonded to the upper and lower channel wall, while leaving open spaces in the channel width for the fluid to flow through. In addition, the long-term hydrophilicity and low nonspecific binding property associated with PEMS surface can be conserved for the nonpatterned area, leading to hydrogel plugs in extremely hydrophilic and permeable environment in a restricted channel space for bubble-free fluid transport and affinity interaction. By immobilization of well-oriented antibodies via protein G on the hydrogel plugs in the channel, estrogen receptor alpha (ERalpha) is demonstrated to be captured quantitatively with high loading capacity and high specificity.

Protein immobilization techniques for microfluidic assays

“…3-D structures have been created by patterning microstructures (e.g., microposts 29,82,83 and micropits 60 ) or through insertion of porous membranes 67,84,85 before assembly of microfluidic chips. In post-assembly approaches, microbeads 54,62,[86][87][88][89][90][91][92][93] can be packed into enclosed channels or various polymers such as hydrogels, [18][19][20][21][22][23][24]57,[94][95][96][97] sol-gels, 64,98,99 polymer monoliths, 61 or membranes 100 can be polymerized in situ. For silica-based 3-D structures such as silica beads 91 and alkoxysilane-based sol-gels, 63 a similar glass/silicon surface immobilization strategy can be used.…”

“…119 Therefore, immobilization methods are generally complex and require multiple steps to implement. 97,120 Because a large numbers of microfluidic devices are made by sealing microchannelpatterned PDMS slabs to glass slides, consideration of glass and PDMS surface properties is often required (e.g., care to avoid nonspecific adsorption to PDMS surfaces when protein is immobilized on glass surface).…”

“…As such, after UV exposure, unreacted benzophenone can be photolyzed again to form a covalent bond. 125 Benzophenone can be also used for free-radical polymerization of hydrogels 97 and surface-grafted polymers.…”

“…Sung et al reported photopatterned 3-D hydrogel plugs for antibody immobilization on a PDMS surface. 97 In order to anchor PA gel to the PDMS (which lacks functional groups), the PDMS was first treated with oxygen plasma. Then, PEI and PAA solutions were injected to yield a multilayer coating on the PDMS surface.…”

Recent Progress of On-line Combination of Preconcentration Device with Microchip Electrophoresis

“…This technique also has been applied to CE and MCE formats, e.g., an application to two-dimensional MCE [23], immunoaffinity gel electrophoresis [24,25], chemical immobilization onto a monolithic column for affinity electrophoretic analysis [26], encapsulation of the biomolecules based on the sol-gel technologies [27], and so on [28], realizing the specific and rapid analyses of the biogenic compounds with small consumptions of samples/reagents. Recently, many researchers focus on the encapsulation of ligands into the hydrophilic gels since the affinity of the encapsulated ligands is well kept as compared to the other chemical immobilization techniques.…”