New Microscope Watches Cells in 3D

Jamie found a story about a new 3D Microscope which creates 3D videos of cells in action. Traditionally scientists have had to choose between high resolution and animation, so no doubt this device will cure the common cold.

Comments

"Cells in action."

[Anonymous Coward]

Sounds hot, hope I can download those videos real soon.

Now I get it.

[tygerstripes]

Ohhhh, I wondered how they did that cool inside-the-body stuff in "House M.D."

Seriously though, this is Big Medicine. I know a couple of guys researching treatments for congenital pancreatic cancer who would kill to get their hands on something like this.

Move over YouTube...

[Delusion_]

...here comes FluTube!

Bad summary...

[ed.mps]

...and bad jokes. A little excerpt for those who didn't RTFA:

It can, for example, capture chromosomes spooling during cell division or a cervical cancer cell shriveling up when treated with acetic acid.

Death by light

[G4from128k]

Although the article does not say so, I'd bet that creatures don't live for very long. All of high-resolution imaging systems that I'm familiar with concentrate so much light on the subject matter that the creature dies within minutes.

Just think of the physics. Most digital sensors need about 10,000 to 100,000 photon to register a full response (i.e, "white") and to see 30 frames per second, that's 300k to 3 million photons per second per pixel. At high resolution a single cell might be 100 pixels by 100 pixels. That means that the poor creature is being hit by 3 to 30 billion photons per second. Even if there's no UV and all heat is removed from the subject, visible light photons in a high enough flux rate will induce various photochemical reactions that damage DNA, denature proteins, and photo-oxidize cellular chemicals. Or to put in another way. consider the amount of light needed to image the average landscape and then concentrate it on a single cell. Even with high-gain amplifiers (= grainy, low-light pictures), the shear concentration of light means the creature doesn't last long.

Re:Death by light

[kebes]

Photo-damage to cells is indeed a concern, but the described technique actually has the advantage that this can minimized as much as physically possible. Many visualization techniques involve either (1) having the cell absorb light, so that you can differentiate different regions based on absorption (may require staining with something sufficiently absorptive), or (2) having something fluoresce, which requires that species to absorb and then re-emit light (typically requires staining or genetic engineering so a target protein is fluorescent). Obviously both (1) and (2) require the sample to actively absorb photos, which means that some amount of photo-heating is unavoidable. Moreover fluorescent molecules often lead to undesired side-reactions and degrade over time (so-called "photo-bleaching"). With fluorescence imaging, you can select an excitation wavelength outside of the absorption bands of everything in solution (especially water!), and thereby minimize photo-heating and photo-damage.

The article says that they are actually imaging the refracted light. Since this technique doesn't require any amount of sample absorption at all, they can use a minimally absorbing wavelength, thereby keeping sample damage to an absolute minimum. In fact since they are measuring refracted light, the technique works best at wavelengths where absorption is as low as possible (but refractive index contrast is as high as possible).

From the description, it doesn't sound like the illumination would be much more intense than what a normal microscope generates. Most cells don't experience significant photo-damage under such illumination conditions.

Some current imaging systems use a raster-scanned focused-laser spot to generate the images. By using high-quality detectors the light-levels can be kept low enough that cell damage is prevented. Thus the technique from the article probably induces less cell damage than currently used techniques. Not to mention that the fact that you don't have to stain or modify the cells eliminates the toxicity (or perturbing effect) or those staining agents.

Re:Death by light

[kebes]

Some more details about the technique. The writeup on the MIT site has more information. The technique is using laser interferometry:

Feld and his colleagues have been able to image live, untreated cells by using an optical technique based on interferometry: a laser beam passed through a sample is compared with a reference beam of similar wavelength that is not passed through the cell. For example, it takes longer for light to travel through a cell than through, say, water. Researchers can measure that time delay, or phase shift, and then can map the cell and its motions on the scale of nanometers.

This appears to be one of the earlier publications on the technique:
"Cellular Organization and Substructure Measured Using Angle-Resolved Low-Coherence Interferometry", Wax A, Yang C, Backman V, Badizadegan K, Boone C, Dasari RR, Feld MS. Biophysical Journal 82: 2256-2264 (2002).

In the experimental section of that article they say:

Broadband light from a superluminescent diode (superluminescent diode (SLD) (EG&G, Gaithersburg, MD), output power 3 mW, center wavelength 845 nm, full width half-maximal bandwidth 22 nm...

This appears to be one of their more recent publications:
"Quantitative phase imaging of live cells using fast Fourier phase microscopy", Niyom Lue, Wonshik Choi, Gabriel Popescu, Takahiro Ikeda, Ramachandra R. Dasari, Kamran Badizadegan, and Michael S. Feld. Applied Optics, Vol. 46, Issue 10, pp. 1836-1842.

In that paper they say:

The second harmonic of the cw Nd:YAG laser (CrytaLaser, special custom-built module; wavelength 532nm, 500 mW) is used as an illumination source for a typical inverted microscope (Axiovert 100, Carl Zeiss).

The illumination sources are not very intense, but are powerful enough to cause cell damage if they were highly focused. From looking over the papers it doesn't seem that this is the case. For what it's worth, the papers do not mention cell damage as being a concern.

Overall the technique seems to have serious promise. It essentially involves doing laser interferometry on the sample at multiple angles, and reconstructing the 3D image. As they mention in their papers, it has the advantage of interfacing with conventional confocal microscope designs. Thus it could be added as an option on existing setups. It appears to have some exacting requirements (like all holography/interferometry it will be sensitive to vibrations, etc.), but overall seems like the type of thing that could be rapidly built into existing labs and commercial instruments.

Press Release Science

[LightPhoenix7]

While interesting, the article had several fallacies in it.

For one, cells can be viewed while alive - fixative isn't always necessary. Motility studies, for exmaple, don't actually kill the cells (or sperm). For another, dyes aren't the only technique to view cells - plasmid insertion into bacteria with a fluorescent marker not only allows cells to be seen, but doesn't harm the cell.

Secondly, I find it decidedly inconvenient that this can only view small images. My current research is in bacterial biofilms - living and dead. I haven't had any trouble viewing living biofilms under a fluorescent or confocal microscope. What if you want to study the chemotaxis of groups of cells? Most cells, eukaryote or prokaryote, talk to each other and can respond differentially to external signals.

Thirdly, even if you can view these cells, only in very specific instances will it give clues about functionality. Sure, that's better than nothing, but it's not the miraculous panacea that the article describes. The mechanics of drug interaction are much more complex than can be determined by simply looking at a cell.

Finally, from a research standpoint, I have to ask how much this costs. Is the cost-benefit ratio really that good that spending large amounts of money to get this is worth it? Especially considering how in reality it has such a limited usage? I would tend to assume no. There may be some very useful things you can do with this, but it just seems like much more of a toy than anything.

Article figure somewhat mislabeled

[Ichoran]

The worm shown in the picture in the article is probably about 200 microns long, not 1 mm--the one shown is recently hatched, not an adult. You can tell because adult worms do not have a pharynx that takes up nearly half the length of the body. Also, in adults you'd be able to see reproductive structures (including eggs).