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A novel method for the preparation of reference free traction force microscopy samples
Traction force microscopy (TFM) is widely used to study mechanotransduction of cellular forces onto their environment. Current methods for sample preparation are time-costly, therefore limiting the feasibility of TFM experiments.
Keywords: Microfabrication, Traction Force Microscopy, Mechanotransduction, Photolithography
Cells transmit mechanical forces onto the extracellular matrix. Changes in such forces occur during growth and development, wound healing and tissue homeostasis, but also in cancer metastasis and disease progression. In order to understand how cellular mechanics respond to changing cues in their environment, it is essential to measure such forces.
A method widely used for this purpose is continuum traction force microscopy with polyacrylamide hydrogels. It relies on the measurement of substrate displacement and consequent calculation of the forces required for such displacements. Typically, fluorescent beads are randomly dispersed in elastic substrates. This allows for high-resolution force maps, but requires the acquisition of an additional reference (load-free) image, which is acquired after cell removal and destruction. This complicates the experimental procedure and does not allow for further experimental steps such as immunofluorescence staining.
Our lab (LTNT) has developed a novel technology for direct, electrohydrodynamically assisted printing of colloidal nanoparticle dispersions to create regular arrays of fluorescent discs. This permits reference-free, high-resolution traction force microscopy.
While very precise, electrohydrodynamic nanodrip printing is a time costly method for sample preparation. This limits the number of samples that can be produced and does not allow to cover larger areas of the samples with arrays. Microfabrication methods offer numerous possibilities to create regular patterns. First steps towards developing a photolithography procedure to pattern substrates have been taken and should be elaborated upon.
Cells transmit mechanical forces onto the extracellular matrix. Changes in such forces occur during growth and development, wound healing and tissue homeostasis, but also in cancer metastasis and disease progression. In order to understand how cellular mechanics respond to changing cues in their environment, it is essential to measure such forces. A method widely used for this purpose is continuum traction force microscopy with polyacrylamide hydrogels. It relies on the measurement of substrate displacement and consequent calculation of the forces required for such displacements. Typically, fluorescent beads are randomly dispersed in elastic substrates. This allows for high-resolution force maps, but requires the acquisition of an additional reference (load-free) image, which is acquired after cell removal and destruction. This complicates the experimental procedure and does not allow for further experimental steps such as immunofluorescence staining. Our lab (LTNT) has developed a novel technology for direct, electrohydrodynamically assisted printing of colloidal nanoparticle dispersions to create regular arrays of fluorescent discs. This permits reference-free, high-resolution traction force microscopy. While very precise, electrohydrodynamic nanodrip printing is a time costly method for sample preparation. This limits the number of samples that can be produced and does not allow to cover larger areas of the samples with arrays. Microfabrication methods offer numerous possibilities to create regular patterns. First steps towards developing a photolithography procedure to pattern substrates have been taken and should be elaborated upon.
The goal of this project is therefore to develop a novel method for patterning hydrogel substrates with fluorescent quantum dot arrays that permit a higher throughput.
The goal of this project is therefore to develop a novel method for patterning hydrogel substrates with fluorescent quantum dot arrays that permit a higher throughput.