Rapid in vitro analysis of force strain in cell signaling pathways for use in tissue engineering
Rapid in vitro analysis of force strain in cell signaling pathways for use in tissue engineering
Facile method allows direct observation of effect of force with much higher throughput than other methods. Other methods to apply force to proteins, such as AFM or optical tweezers, are limited by the difficulty of setting up experiments and the time to complete an analysis. Additionally, these instruments are very expensive and difficult to operate and maintain. This method allows for multiple measurements to be conducted at the same t...
New York, NY, United States
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Background 

Cells are cultured on a flexible silicon substrate, and treated with a detergent. This treatment dissolves the majority of the cell, but leaves behind the bare cytoskeleton proteins which remain bound to the substrate. The silicon can be stretched or compressed to directly apply force to the cytoskeletal structures. This allows for rapid in vitro analysis of these structures under a force load. For example, the binding of labeled proteins can be monitored to determine the effect of contraction or stretching on binding. Furthermore, this technique can be conducted in the presence of potential drugs, to quickly screen for their effect and potency in modulating tissue function or stiffness. This application is highly sought after for the development of engineered tissues, as well as novel treatments for various injuries and tissue damage.

Facile method allows direct observation of effect of force with much higher throughput than other methods

Other methods to apply force to proteins, such as AFM or optical tweezers, are limited by the difficulty of setting up experiments and the time to complete an analysis. Additionally, these instruments are very expensive and difficult to operate and maintain. This method allows for multiple measurements to be conducted at the same time, greatly increasing the rate of experiments that can be conducted by a single researcher. The speed of analysis will be greatly useful for high-throughput screening of small molecules to assess their medical use in tissue engineering and injury treatment.

This method was verified by correctly identifying signaling molecules that bind only to force strained cytoskeletal proteins.

Lead Inventor:

Michael Sheetz, Ph.D.

Applications:

  • Investigate the effect of a force load on the cytoskeleton
  • Monitor binding of other proteins, and the dependency of that binding upon force
  • Test the effects of potential drugs in tissue engineering or injury treatment
  • Culture cells in conditions that resemble a strained extracellular matrix

Advantages:

  • Significantly cheaper and faster than other methods to apply force to cytoskeletal structures
  • Versatile applications in biomedical research from tissue engineering to researching mechanotransduction
  • High-throughput; one researcher can cover multiple experiments or test multiple drugs in a short time
  • Doesn’t require advanced and specialized instruments to operate
  • Cytoskeletons are obtained by treating cells; protein purification and reconstitution is not required
  • Silicon substrate with matrix molecules more closely resembles biological tissue than most experimental substrates

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