Molecular Imaging of a Micropatterned Biological Ligand on an Activated Polymer Surface (original) (raw)
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Microstamping on an activated polymer surface (MAPS) is a methodology that enables biomolecules to be patterned on polymers with micrometer spatial resolution. MAPS combines homogeneous surface derivatization of a polymer to introduce a reactive functional group followed by reactive microcontact printing (µCP) of a biological ligand of interest, linked to an appropriate reactive group. We demonstrate here that polyethylene, polystyrene, poly(methyl methacrylate), and poly(ethylene terephthalate) films can be successfully modified to introduce COOH groups on their surfaces, which can be subsequently patterned by reactive µCP of amine-terminated biotin after derivatization of the COOH groups with pentafluorophenol. X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry (TOF-SIMS) confirmed the chemistry of MAPS at each stage of the derivatization of the polymer surfaces and reactive µCP of biotin. Micropatterned biotin surfaces fabricated by MAPS were patterned with streptavidin by exploiting molecular recognition between biotin and streptavidin. The formation of streptavidin patterns was examined by fluorescence microscopy of Alexa488-labeled streptavidin and by TOF-SIMS imaging of 15 N-labeled recombinant streptavidin, bound to biotin patterns. The contrast in the streptavidin micropatterns was optimized by examining the effect of blocking agents and streptavidin incubation time. Maximum contrast was obtained for binding of 0.1µM streptavidin from a buffer containing 0.02% (v/v) Tween 20 detergent for an incubation time of 1 min.
2023
Rough, capillary-active surfaces remain demanding substrates for microcontact printing (µCP), as the diffusive mobility of the ink thereon drastically limits the printing resolution. To reduce ink smearing, we developed a polymer-supported μCP, which includes a stamp with a polymer brush-decorated surface. The ink molecules are thereby bound into the stamp-bound brush matrix, from where they may be transferred to the substrate, which exclusively occurs during the contact of both interfaces. Conventionally, Slygard184-based polydimethylsiloxane (PDMS) stamps are used for µCP. The material's surface must be functionalized in a multi-step procedure for the protocol. In addition, Sylgard comes along with a drawback of a persistent leakage oligomeric PDMS (oPDMS), which can contaminate the substrate. To circumvent these problems, we developed a novel stamp material, that (i) enables a straightforward polymer grafting, and (ii) shows a low tendency of oPDMS leakage. We prepare the stamp with a commercially available amino-functional PDMS prepolymer, and a polymer-ic crosslinker that can be used for a controlled photoiniferter reversible addition and fragmentation chain transfer (PI-RAFT) polymerization. The prepared stamp shows elastic properties at the relevant strain region, is compatible with brush formation, and has been demonstrated demonstrated suitable to transfer precise patterns on rough capillary-active oxide surfaces. ASSOCIATED CONTENT SUPPORTING INFORMATION NMR spectra, SEC data, mechanical data, light and fluorescence microscopy images, AFM height images of samples are found in the supporting information.
Journal of Colloid and Interface Science, 2010
Time-of-Flight Secondary Ion Mass Spectrometry (TOF-SIMS) was applied to validate GRGDS peptide patterned surfaces. The structuring of the surfaces included several steps: micro contact printing (lCP), chemical etching and aminofunctionalization followed by chemical coupling of spacer-linked GRGDS peptides via an isothiocyanate anchor. TOF-SIMS analysis of characteristic ions and molecular fragments with a lateral resolution of 100 nm allowed proving the change in chemical properties of the surface with each step during the structuring process. We found that the application of polydimethylsiloxane as stamp material resulted in the contamination of the surface with this polymer. TOF-SIMS investigations, however, also showed that during the preparation process the contaminations were removed and do not influence the bio functionality of the surface patterns. The results of the surface analysis carried out with TOF-SIMS were confirmed by complementary cell adhesion experiments with murine fibroblasts. As a result, specific cell adhesion restricted to GRGDS peptide functionalized areas was obvious by the formation of focal adhesion contacts in the fibroblasts. Thus, TOF-SIMS is the method of choice in chemical characterization of surfaces in structuring and functionalization processes, because it offers the opportunity to follow surface contamination during the preparation process and to assess the influence of the contamination on the applicability of the final substrate.
ToF-SIMS multivariate analysis of surface-grafted small bioactive molecules
Biointerphases, 2015
In the development of bioactive coatings on biomaterials, it is essential to characterize the successful fabrication and the uniformity of intended coatings by sensitive surface analytical techniques, so as to ensure reliable interpretation of observed biointerfacial responses. This can, however, be challenging when small bioactive molecules are grafted onto biomaterials surfaces at sub- and near-monolayer densities. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) provides the required sensitivity, but ion signals from small grafted molecules may still be dominated by fragment ions from the underlying polymer. In such cases, multivariate analysis provides valuable enhancement of spectral data, as illustrated here by examples comprising the surface grafting of bioactive serrulatane molecules, the peptide GRGDSP, the oligonucleotide 15-thymidine, and the antifungal compound Amphotericin B. The authors also show how ToF-SIMS plus principal component analysis can distinguish b...
Mask and Lithography …, 2008
Top surface imaging (TSI) techniques using vapor or liquid phase silylation have been investigated extensively as alternatives to conventional resist processing. However, earlier imaging schemes such as diffusion enhanced silylated resist (DESIRE) and digital top surface imaging showed several difficulties limiting the successful application of such TSI approaches. In the case of DESIRE, additional CF 4 plasma descum process was required to remove the thin layer of Si incorporated into the cross-linked regions, as some of the Si remained even in the unexposed regions. Also, difference in the cross-linking density and subsequent amount of silicon incorporation across the width of an optically projection printed feature led to non-uniform silylation profiles resulting the difficulty with critical dimension (CD) control of the feature and increased the LER of the overall process. In the case of digital TSI, even though it was developed to overcome these problems with the cross-linking-based silylation process, the concentration of active sites in the exposed polymer varies across the feature width due to the non-uniform energy deposition profile across a feature which results from the non-ideal aerial image produced using optical projection tools. In this study, we have used a diazoketo-functionalized polymer as the platform for the immobilization of aminefunctionalized poly(dimethyl siloxane) (amine-PDMS). The diazoketo functional groups undergo Wolff rearrangement to generate carboxylic acid groups upon UV light exposure. This chemistry is exploited to create alternate hydrophilic/hydrophobic patterned regions by selective UV light exposure. The hydrophilic regions that contain carboxylic acid groups predominantly are further used to immobilize amine-PDMS by amide bond formation using carbodiimide coupling chemistry. Due to the high silicon content, the immobilized PDMS acts as the etch barrier for the subsequent oxygen plasma reactive ion etching (O 2-RIE) process. Thus, a negative-tone pattern has been successfully generated using O 2-RIE process. An amine-PDMS with a molecular weight of 900 was used in this study. Auger electron spectroscopy was employed to characterize the immobilization of amine-PDMS onto UV light exposed regions of diazoketo-functionalized polymer surface. Atomic force microscopy was used to study the surface smoothness after O 2-RIE process. Scanning electron microscopy was used to image the pattern profiles formed after O 2-RIE process. High resolution pattern profiles are obtained using the TSI process reported in this study.
Langmuir, 1999
A patterned array consisting of a microstructured layer of octadecyltrichlorosilane (OTS) and (3-((2,4dinitrophenyl)amino)propyl)triethoxysilane (DNP analogue) was assembled onto a Si wafer using the microcontact printing method. The microstructured, patterned support was imaged by AFM, using a bare Au tip, a hydrophobic alkyl mercaptan-functionalized Au tip, and a hydrophilic hydroxyalkyl mercaptanmodified Au tip. The lateral friction forces between the tip and the patterned surface are controlled by hydrophobic-hydrophobic and hydrophobic-hydrophilic interactions and eventually H bonds between the tip and the functionalized surface. The (3-((2,4-dinitrophenyl)amino)propyl)triethoxysilane domains of the microstructured surface act as antigens for the anti-dinitrophenyl antibody (anti-DNP-Ab). Interaction of the patterned support with the DNP-Ab solution yields an overall hydrophilic interface due to the association of the antibody to the entire support. Analysis of the adhesive and friction interactions between the Au tip and the DNP-Ab associated with the OTS and DNP analogue regions, and the roughness factors of the respective domains that include the DNP-Ab, enables us to conclude that the DNP-Ab associates nonspecifically to the OTS sites, while specific binding of the DNP-Ab occurs on the DNP antigen regions.
Patterning small-molecule biocapture surfaces: microcontact insertion printing vs. photolithography
Chemical Communications, 2011
oxaundecan-11-biotin (BEG) was purchased from ProChimia (Sopot, Poland). N-Hydroxysuccinimide (NHS), N-ethyl-N-(dimethylaminopropyl)-carbodiimide (EDC), 5-hydroxytryptamine hydrochloride (5-HT), 3,4-hydroxytyramine hydrochloride (3,4-dihydroxyphenylethylamine; dopamine; DA), and bovine serum albumin (BSA) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Commercial grade ethanol (EtOH) was from Pharmaco-AAPER (Brookfield, CT, USA). Rabbit anti-serotonin polyclonal antibodies and rabbit anti-dopamine polyclonal antibodies were procured from Millipore (Temecula, CA, USA). AlexaFluor 488-labeled goat anti-rabbit antibodies (absorbance max at 495 nm, emission max at 519 nm), AlexaFluor 546-labeled goat anti-rabbit antibodies (absorbance max at 556 nm, emission max at 573 nm), streptavidin, and AlexaFluor 488-tagged streptravidin (absorbance max at 494 nm, emission max at 521 nm) were purchased from Materials and Methods
Japanese Journal of Applied Physics, 2008
Patterning biomolecules at the micron or nano-scale presents a major challenge for the elaboration of integrated biochips. Among the different techniques that are emerging, Microcontact printing (mCP) appears as one of the most promising due to its simplicity and low cost. However, mCP exhibits a severe limitation because only one type of molecule can be deposited at a time using a poly(dimethyldiloxane) (PDMS) stamp. We present a new process called one-step-multiple-mCP (OSM-mCP) using a specific multilevel PDMS stamp which allows the patterning of two different molecules in one step. Our method based on the elastomeric properties of a PDMS stamp can print self aligned patterns of two different molecules by pressuring homogenously the top side of the stamp. The stamp levels inked by two different molecules contact the surface sequentially. OSM-mCP of two proteins, here bovin serum albumin BSA and streptavidin is demonstrated and this method can be applied to others couples of molecules. These results show that patterning biomolecules with OSM-mCP process opens new perspectives for soft lithography by enabling the fabrication of complex patterns of different molecules.