Kanazawa University NanoLSI Podcast:Endoscopy of a Living Cell on the Nanoscale

Endoscopy of a living cell on the nanoscale

 Transcript of this podcast

 Hello and welcome to the NanoLSI podcast. Thank you for joining us today. In this episode we feature the latest research by Takeshi Fukuma at the Kanazawa University NanoLSI.

 The research described in this podcast was published in Science Advances in December 2021

 Kanazawa University NanoLSI website

https://nanolsi.kanazawa-u.ac.jp/en/

Endoscopy of a living cell on the nanoscale

Researchers at Kanazawa University report in Science Advances a new technique for visualizing the inside of a biological cell. The method is an extension of atomic force microscopy and offers the promise of studying nanoscale inner cell dynamics at high resolution in a non-destructive way.

In order to advance our understanding of how biological cells function, visualizing the dynamics of intra-cellular components on the nanoscale is of key importance.  Current techniques for imaging such dynamics are not optimal — for example, fluorescence microscopy can visualize ‘labeled’ molecules but not the target components themselves.  Now Takeshi Fukuma from Kanazawa University and his colleagues have developed a label-free, non-destructive nanoimaging method, which they call nanoendoscopy-AFM – it’s a version of atomic-force microscopy that can be deployed within a living cell.  The research was carried out as a collaboration between Kanazawa University and the National Institute of Advanced Industrial Science and Technology (AIST), with Marcos Penedo, the lead author of the publication reporting the new method, recently moving from Kanazawa University’s Nano Life Science Institute (WPI-NanoLSI) to the École Polytechnique Fédérale de Lausanne, Switzerland.

So what is AFM in the first place?

The principle of AFM – or atomic force microscopy ti give its full title - is to have a very small tip move over the surface of a sample.  During this ‘xy’ scanning motion, the tip, attached to a small cantilever, will follow the sample’s height, that is, the (‘z’) dimension or profile, producing a measurable force on the cantilever.  The magnitude of the force can be back-converted into a height value; the resulting height map provides structural information about the sample’s surface.

The researchers designed a novel AFM setup where the needle-like tip is brought in and out of the interior of a cell.  The process is reminiscent of an endoscopy — the procedure of looking at an organ from the inside, by inserting a small camera attached to a thin tube into the body — which is why Fukuma and colleagues call their technique nanoendoscopy-AFM.  Letting the nanoneedle travel in an ‘xyz’ trajectory, and going in and out of the cell results in a 3D map of its structure.  They tested the technique on a cell from the so-called HeLa cell line commonly used in medical research, and could clearly identify internal granular structures in a scanned volume of 10 x 10 x 6 µm3.

But how does the cell fare under this kind of interrogation?

During a scan, the nanoneedle penetrates the cell membrane (and the nuclear membrane) many times.  The scientists checked whether this repeated penetration causes any damage to the cell.  They performed a viability test on HeLa cells by using two fluorescent marker molecules.  One molecule emits green fluorescence from a living cell, the other red fluorescence from (the nucleus of) a dead cell.  The researchers found that when using nanoprobes smaller than 200 nm, nanoendoscopy-AFM does not severely damage cells.

The method is also particularly useful for probing surfaces within the cell, for example the inner side of the cell membrane or the surface of the cell nucleus.  Fukuma and colleagues call this application 2D

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