Editorial Feature

New Tissue Expansion Technique for High Resolution Microscopy Imaging

Researchers from the Massachusetts Institute of Technology (MIT) Department of Brain and Cognitive Sciences have recently developed a new way to obtain high-resolution images of tissue samples using conventional light microscopes.

Most importantly, the high-resolution images obtained by this new technique are possible at a fraction of the cost of other techniques that offer a similar resolution7.

In 2015, Edward Boyden’s team developed a method called “expansion microscopy” to explore a new possibility of expanding the tissue sample before imaging it with a conventional light microscope, as opposed to simply magnifying the image by use of the lenses of the microscope.

The tissue samples were embedded in a dense, evenly generated crosslinked gel made of a very absorbent polymer material that is also used in diapers called “polyacrylate.”  “Barcodes” bearing antibodies that bind to specific targets were used to target the cellular proteins of interest. While simultaneously using cross-linking molecules, polymers that make up the expandable gel are attached to the barcodes, which are made up of DNA that are present on the antibodies7.

As the gel swells by absorbing water, the proteins that normally hold the tissues together break down and expand away from each other, resulting in a complete enlargement of the tissue sample7. Fluorescent probes that bind to the DNA barcodes can also be used to label the enlarged sample, allowing for the imaging by use of commercially available microscopes, whose resolution is usually limited to hundreds of nanometers7.

In the recent article published in Nature Methods on April 17th, 2017, Boyden’s team showed that resolutions of about 25 nm is possible by expanding the tissue sample for a second time before the imaging takes place7. To improve the resolution further, while also preserving the stability of the tissue sample, the researchers used a new technique called “iterative expansion.”Boyden’s team of researchers showed that tissue samples can be expanded up to 100-fold by reducing the number of cross linking molecules that hold the polymer together, resulting in an image resolution of about 60 nanometers (nm) 7. However, this reduction in the density of cross-linkers affected the organization of the polymers, which resulted in decreased tissue stability and resolution loss.

In this expansion method, a new gel is used to increase the size of the tissue for a second time after the first tissue expansion, thereby enlarging the tissue in three dimensions and preventing any loss of resolution7.

Boyden’s team used their technique to image synapses, which are neuronal junctions through which nerve cells, or neurons, communicate with each other7. While the researchers’ earlier expansion microscopy technique allowed them to image scaffolding proteins that help in organizing hundreds of other proteins that are found in synapses, the increased resolution of the iterative expansion technique has now allowed them to visualize finer details of the tissue sample, such as receptors that are located on the surface of the postsynaptic neuron7.

Using a technique called temporal multiplexing, which involves the labeling of different molecules with a specific fluorescent probe of a distinct color one at a time after taking a picture, and subsequent washing before the next fluorophore labelling, it is possible to achieve nanoscale-resolution imaging over three dimensional (3D) volumes7.

While the images produced by the iterative expansion technique cannot match the resolution of TEM images yet, the 25-nm resolution obtained by this method is similar to that of the STORM. The iterative expansion, however, is rather simple to perform, with no requirement of any specialized equipment or chemicals. The technique is also extremely inexpensive as compared to other microscopy methods of this magnitude.

While remaining compatible with large-scale 3D imaging, the rapid rate of imaging production of this technique is also particularly striking7. This group of MIT researchers is hopeful that they could potentially map out the organization of various scaffolding and signaling proteins located at the synapse by combining temporal multiplexing with this iterative expansion technique.

Boyden’s team is currently investigating the possibility of incorporating a third round of expansion in order to achieve a resolution of about 5 nm7.


  1. “Medical Definition of Microscopy” – MedicineNet.com
  2. “Light Microscopy” – Rice University
  3. “Scanning Electron Microscope” – Purdue University
  4. “Correlated fluorescence and 3D electron microscopy with high sensitivity and spatial precision.” W. Kukulski, M. Schorb, et al. Journal of Cell Biology. 2011. DOI: 10.1083/jcb.201009037.
  5. “Scanning Electron Microscopy” – Microscope Master
  6. “Transmission Electron Microscope (TEM)” – Encyclopedia Britannica
  7. “High-resolution imaging with conventional microscopes” – MIT News
  8. “Super-resolution Microscopy” – University of California San Francisco
  9. Image Credit: Shutterstock.com/SydaProductions

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Benedette Cuffari

Written by

Benedette Cuffari

After completing her Bachelor of Science in Toxicology with two minors in Spanish and Chemistry in 2016, Benedette continued her studies to complete her Master of Science in Toxicology in May of 2018. During graduate school, Benedette investigated the dermatotoxicity of mechlorethamine and bendamustine; two nitrogen mustard alkylating agents that are used in anticancer therapy.


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