Chemoselective Immobilization of Proteins by Microcontact Printing and Bio-orthogonal Click Reactions (original) (raw)
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Light-Activated Affinity Micropatterning of Proteins on Self-Assembled Monolayers on Gold
We describe a method to pattern proteins onto a photolabile "caged" biotin-derivatized self-assembled monolayer (SAM) on gold, which we term light-activated affinity micropatterning of proteins (LAMP). LAMP is a multistep patterning process with considerable flexibility in its implementation. First, a reactive SAM on gold is formed from a mixture of 11-mercaptoundecanol and 16-mercaptohexadecanoic acid. Next, the carboxylic acid end groups in the SAM are coupled to methyl R-nitropiperonyloxycarbonyl biotin succinimidyl ester (caged biotin ester) through a diamine linker. The caged biotin is then deprotected in regions irradiated by masked UV light, and subsequent incubation with streptavidin results in selective binding of streptavidin to the irradiated regions. Micropatterning of various proteins has been demonstrated with a spatial resolution of ∼6 µm by confocal microscopic imaging of fluorophore-labeled proteins, and a contrast ratio of ∼4:1 was determined by direct ellipsometric imaging of streptavidin. Immobilization of biotinylated antibodies on the streptavidin pattern indicates that LAMP can enable spatially resolved micropatterning of different biomolecules by repeated cycles of spatially defined photodeprotection of biotin, streptavidin incubation, followed by immobilization of the biotinylated moiety of interest.
Journal of materials science. Materials in medicine, 1999
Micrometer-scale patterns of a defined surface chemistry and structure were produced on both ultraflat Au(1 1 1) and on gold-coated monocrystalline silicon surfaces by a method combining microcontact printing, wet chemical etching and the replacement of etch-resist self-assembled monolayers (SAMs) by functionalized or reactive SAMs. Key steps in this methodology were characterized by X-ray photoelectron spectroscopy (XPS), ellipsometry and contact angle measurements. The covalent immobilization of (functional) biological systems on these surfaces was tested using an N-hydroxysuccinimide ester omega-functionalized disulphide (DSU), which covalently binds primary amines without the need for further activation steps. Atomic force microscope images of native collagen V single molecules immobilized on these patterned surfaces revealed both high spatial resolution and strong attachment to the monolayer/gold surface. Microcontact printing of DSU is shown to be feasible on specially prepare...
Biomedical Microdevices - BIOMED MICRODEVICES, 2001
Two-dimensional protein micropatterning with immobil-ization of IgG and poly (ethylene glycol) (PEG) on patterned Au and Si surfaces was performed through a new technique. The technique for micropatterning is based on a chemical selectivity method by creating chemical bonding between protein, self-assembled monolayers (SAMs) and substrates rather than physical means. The substrates used in this study are pre-fabricated with silicon wafer patterned with arrays of gold squares. The silicon regions of the substrate are modified with polyethylene glycol (PEG) to resist protein adsorption and cell adhesion. The gold regions on the substrate are first immobilized with bifunctional SAM layers that can covalently bound adhesion proteins for individual cell attachment against a PEG background. The surface coatings are characterized by contact angle measurement, ellips-ometry, and atomic force microscopy (AFM). The patterns of fluorescence-labeled proteins are examined using fluorescence micr...
Photocatalytic Nanolithography of Self-Assembled Monolayers and Proteins
N anofabrication is a key underpin-11 ning technology in nanoscience. 12 Self-assembled monolayers (SAMs) 1À4 13 provide a powerful and versatile means by 14 which to control interfacial molecular struc-15 ture and interactions, and the modification 16 of SAMs at nanometer length scales has 17 attracted widespread attention during the 18 past two decades. 5 A variety of approaches 19 has been developed, including dip-pen 20 nanolithography, 6À12 nanoshaving, and 21 nanografting, 13À15 local oxidation, 16,17 elec-22 tron beam lithography, 18À21 microcontact 23 printing, 22À24 and scanning near-field litho-24 graphy (SNP), 25À29 all of which have been 25 employed successfully to pattern SAMs at 26 better than 100 nm resolution. 27 The control of protein organization on 28 sub-100 nm length scales presents particu-29 lar challenges. 9,20,23,30À40 Proteins are able 30 to adsorb strongly to most surfaces, because 31 they display a wide variety of functional 32 groups (charged and uncharged, cationic 33 and anionic, hydrophilic and hydrophobic) 34 and because they exhibit substantial con-35 formational freedom. The first requirement 36 for protein patterning is thus an effective 37 means to control nonspecific adsorption. A 38 number of approaches have been reported 39 in the literature, but the most widely used ones 40 have been based around poly(ethylene 41 glycol) 41 and its derivatives, including 42 oligo(ethylene glycol) (OEG)-terminated 43 SAMs, 42À48 plasma-polymerized films, 49À51 44 and poly(oligoethylene glycol methacrylate) 45 brushes. 52À54 46 One approach to protein patterning is to 47 form a protein-resistant surface and then 48 selectively introduce protein-binding re-49 gions. For example, protein-resistant OEG-50 functionalized nitrophenylethoxycarbonyl-51 protected silane films have been selectively 52 deprotected by near-field optical methods 53
Protein-Resistant Self-Assembled Monolayers on Gold with Latent Aldehyde Functions
Langmuir, 2007
In the present study, oligo(ethylene glycol) (OEG)-linked alkanethiols were synthesized which carry a vicinal diol on one end of the OEG chain. After self-assembled monolayer (SAM) formation on gold, the vicinal diols were converted into aldehyde functions by exposure to aqueous NaIO 4 , as previously used for SAMs with OEG chains buried in the center of the SAM [Jang et al. Nano Lett. 2003, 3, 691-694]. Mixed SAMs with latent aldehydes on 5% of the OEG termini showed high protein resistance, which greatly slowed the kinetics of protein coupling on the time scale of minutes. Small bioligands (such as biocytin hydrazide) or small heterobifunctional crosslinkers (maleimidopropionyl hydrazide, pyridyldithiopropionyl hydrazide) with hydrazide functions were efficiently bound to the aldehyde functions on the SAM, providing for specific capture of streptavidin or for fast covalent binding of proteins with free thiols or maleimide functions, respectively. In conclusion, OEG-terminated SAMs with latent aldehydes serve as protein-resistant sensor surfaces which are easily functionalized with small ligands or with heterobifunctional crosslinkers to which the bait molecule is attached in a subsequent step.
Langmuir, 2010
To study complex cell behavior on model surfaces requires biospecific interactions between the interfacing cell and material. Developing strategies to pattern well-defined molecular gradients on surfaces is difficult but critical for studying cell adhesion, polarization, and directed cell migration. We introduce a new strategy, microfluidic SPREAD (Solute PeRmeation Enhancement And Diffusion) for inking poly(dimethylsiloxane) (PDMS) microfluidic cassettes with a gradient of alkanethiol. Using SPREAD, an oxyamine-terminated alkanethiol is able to permeate into a PDMS microfluidic cassette, creating a chemical gradient, which can subsequently be transfer printed onto a gold surface to form the corresponding chemoselective gradient of oxyamine-alkanethiol self-assembled monolayer (SAM). By first patterning regions of the gold surface with a protective SAM using microfluidic lithography, directional gradients can be stamped exclusively onto unprotected bare gold regions to form single cell gradient microarrays. The microfluidic SPREAD strategy can also be extended to print micrometer-sized islands of radial SAM gradients with excellent geometric resolution. The immobilization of a cell adhesive Arg-Gly-Asp (RGD)-ketone peptide to the SPREAD stamped oxyamine-alkanethiol SAMs provides a stable interfacial oxime linkage for biospecific studies of cell adhesion, polarity, and migration.
Experimental Cell Research, 1997
This paper describes a convenient methodology for monolayer (SAM) of alkanethiolates [1-3]. This work patterning substrates for cell culture that allows the uses microcontact printing (mCP) [4, 5] to pattern SAMs positions and dimensions of attached cells to be coninto regions terminated in methyl groups and tri(ethyltrolled. The method uses self-assembled monolayers ene glycol) groups; SAMs terminated in methyl groups (SAMs) of terminally substituted alkanethiolates promote the hydrophobic adsorption of protein and (R(CH 2) 11-15 S0) adsorbed on optically transparent SAMs terminated in oligo(ethylene glycol) groups resist films of gold or silver to control the properties of essentially entirely the adsorption of protein. Immerthe substrates. SAMs terminated in methyl groups sion of these patterned substrates in a solution conadsorb protein and SAMs terminated in oligo(ethytaining fibronectin results in adsorption of protein spelene glycol) groups resist entirely the adsorption of cifically on the methyl-terminated regions of the SAM; protein. This methodology uses microcontact printsubsequent placement of these substrates in medium ing (mCP)-an experimentally simple, nonphotocontaining a suspension of bovine capillary endothelial lithographic process-to pattern the formation of (BCE) cells results in the attachment and spreading of SAMs at the micrometer scale; mCP uses an elastocells predominantly on the regions of the patterned meric stamp having at its surface a pattern in relief SAM that present fibronectin. to transfer an alkanethiol to a surface of gold or silver in the same pattern. Patterned SAMs having hy-Cell adhesion is important for control of cell shape, drophobic, methyl-terminated lines 10, 30, 60, and 90 growth, and function. Although it is not yet routine to mm in width and separated by protein-resistant reexperimentally vary the adhesion of cells in a congions 120 mm in width were prepared and coated trolled manner, several groups have developed methwith fibronectin; the protein adsorbed only to the ods to control spatially the attachment of cells to submethyl-terminated regions. Bovine capillary endostrates [6-20]. Much of this work has employed glass thelial cells attached only to the fibronectin-coated, slides that were modified with monolayers of alkylsiloxmethyl-terminated regions of the patterned SAMs. anes and has used photolithographic methods to pat-The cells remained attached to the SAMs and contern these monolayers. For example, Kleinfeld and cofined to the pattern of underlying SAMs for at least workers patterned quartz substrates into alkylsilox-5-7 days. Because the substrates are optically transanes terminated in methyl groups and amino groups parent, cells could be visualized by inverted microswith sizes of features down to 5 mm [9]. When plated copy and by fluorescence microscopy after fixing and in the presence of serum, cerebellar cells attached only staining with fluorescein-labeled phalloidin. ᭧ 1997 to the amino-terminated regions of the substrate; we Academic Press presume that proteins of the serum that do not support cell attachment adsorbed to the methyl-terminated regions. Though this and much other work has provided