Photobleaching-activated micropatterning on self-assembled monolayers

Scrimgeour, Jan; Kodali, Vamsi; Kovari, Daniel T.; Curtis, Jennifer E.

doi:10.1088/0953-8984/22/19/194103

Cited by 10 publications

(12 citation statements)

References 23 publications

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“…The study and control of such processes necessarily rely on the fabrication of in-vitro models mimicking the complexity of the extracellular environment. With this goal, several biopatterning techniques have been developed during the past decades, including microdispensing techniques [10] and microfluidic systems [11], optical lithography [12,13], photochemical patterning [14][15][16]. Nowadays, the most common technique is microcontact printing [17][18][19][20], which involves the creation of a stamp by soft-lithography from a microfabricated mold, and the subsequent transfer of molecules from the stamp to the final substrate with a fast and inexpensive printing process.…”

Section: Introductionmentioning

confidence: 99%

Thermochemical scanning probe lithography of protein gradients at the nanoscale

Albisetti

Carroll

et al. 2016

Nanotechnology

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Patterning nanoscale protein gradients is crucial for studying a variety of cellular processes in vitro. Despite the recent development in nano-fabrication technology, combining nanometric resolution and fine control of protein concentrations is still an open challenge. Here, we demonstrate the use of thermochemical scanning probe lithography (tc-SPL) for defining micro- and nano-sized patterns with precisely controlled protein concentration. First, tc-SPL is performed by scanning a heatable atomic force microscopy tip on a polymeric substrate, for locally exposing reactive amino groups on the surface, then the substrate is functionalized with streptavidin and laminin proteins. We show, by fluorescence microscopy on the patterned gradients, that it is possible to precisely tune the concentration of the immobilized proteins by varying the patterning parameters during tc-SPL. This paves the way to the use of tc-SPL for defining protein gradients at the nanoscale, to be used as chemical cues e.g. for studying and regulating cellular processes in vitro.

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Section: Introductionmentioning

confidence: 99%

Thermochemical scanning probe lithography of protein gradients at the nanoscale

Albisetti

Carroll

et al. 2016

Nanotechnology

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“…Likewise, continuous gradients (Figure 4c) show an increasing intensity signal for linearly increasing pixel intensity values with a slight saturation towards higher intensity values (Figure 4d). We assume that depletion of fluorophores in regions with intense irradiation is responsible for the saturation effects seen for high intensity values, as has been previously suggested 24…”

Section: Resultsmentioning

confidence: 87%

“…In this study, fluorescence intensity values are taken as a measure for the amount of bound streptavidin, as is common practice in protein immobilization 15, 21, 22, 24, 26. As described in the literature, protein density has been quantified using dried fluorescently labeled proteins and radioactively labeled proteins, thereby demonstrating that fluorescence intensity can indeed be taken as a measure of protein loading 22, 31.…”

Section: Resultsmentioning

confidence: 99%

“…Upon irradiation an oxygen‐dependent energy‐transfer process transforms the fluorophore into a free‐radical species that binds to the protein and thereby immobilizes the attached ligand 23. This coupling strategy has also been used to couple ligands to unsaturated compounds such as methacrylate silanes instead of BSA 24. The main advantage of PAP compared to conventional photochemical or photolithographic techniques is that the fluorophore can be chosen from a wide selection of available dyes, so that the process can be carried out at wavelengths that are not harmful to proteins.…”

Section: Introductionmentioning

confidence: 99%

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Maskless Projection Lithography for the Fast and Flexible Generation of Grayscale Protein Patterns

Waldbaur¹,

Waterkotte²,

Schmitz

et al. 2012

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Protein patterns of different shapes and densities are useful tools for studies of cell behavior and to create biomaterials that induce specific cellular responses. Up to now the dominant techniques for creating protein patterns are mostly based on serial writing processes or require templates such as photomasks or elastomer stamps. Only a few of these techniques permit the creation of grayscale patterns. Herein, the development of a lithography system using a digital mirror device which allows fast patterning of proteins by immobilizing fluorescently labeled molecules via photobleaching is reported. Grayscale patterns of biotin with pixel sizes in the range of 2.5 μm are generated within 10 s of exposure on an area of about 5 mm(2) . This maskless projection lithography method permits the rapid and inexpensive generation of protein patterns definable by any user-defined grayscale digital image on substrate areas in the mm(2) to cm(2) range.

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“…Later Schwarz et al introduced a new method to compute traction forces only at the focal adhesion site of the cell by assuming that the cell force transfer occurs only through these sites[50]. Some novel platforms, such as the photobleaching-activated monolayer with adhesive micro-patterns developed by Scrimgeour et al [64] and the elastic substrates with micro-contact printing demonstrated by Stricker et al [65], were also used to characterize the cell traction force. Furthermore, a FEA-based technique was also developed by Yang et al to greatly improve the accuracy of traction force calculations [56].…”

Section: Introductionmentioning

confidence: 99%

A Novel Cell Traction Force Microscopy to Study Multi-Cellular System

et al. 2014

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Traction forces exerted by adherent cells on their microenvironment can mediate many critical cellular functions. Accurate quantification of these forces is essential for mechanistic understanding of mechanotransduction. However, most existing methods of quantifying cellular forces are limited to single cells in isolation, whereas most physiological processes are inherently multi-cellular in nature where cell-cell and cell-microenvironment interactions determine the emergent properties of cell clusters. In the present study, a robust finite-element-method-based cell traction force microscopy technique is developed to estimate the traction forces produced by multiple isolated cells as well as cell clusters on soft substrates. The method accounts for the finite thickness of the substrate. Hence, cell cluster size can be larger than substrate thickness. The method allows computing the traction field from the substrate displacements within the cells' and clusters' boundaries. The displacement data outside these boundaries are not necessary. The utility of the method is demonstrated by computing the traction generated by multiple monkey kidney fibroblasts (MKF) and human colon cancerous (HCT-8) cells in close proximity, as well as by large clusters. It is found that cells act as individual contractile groups within clusters for generating traction. There may be multiple of such groups in the cluster, or the entire cluster may behave a single group. Individual cells do not form dipoles, but serve as a conduit of force (transmission lines) over long distances in the cluster. The cell-cell force can be either tensile or compressive depending on the cell-microenvironment interactions.

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Photobleaching-activated micropatterning on self-assembled monolayers

Cited by 10 publications

References 23 publications

Thermochemical scanning probe lithography of protein gradients at the nanoscale

Thermochemical scanning probe lithography of protein gradients at the nanoscale

Maskless Projection Lithography for the Fast and Flexible Generation of Grayscale Protein Patterns

A Novel Cell Traction Force Microscopy to Study Multi-Cellular System

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