A nanoelectronics-blood-based diagnostic biomarker for ME/CFS (2019) Esfandyarpour, Davis et al

"The impedance results are integrations of the impedance that are attributed to the media− sensor surface interactions (Zm-s), cell−cell interactions (Zc-c), cell−sensor surface adhesion (Zc-s), impedance of cells (membrane capacitance Cm, cytoplasm conductivity of the cells, σcp), resistance of the solution (Rs), and other components (e.g., proteins, exosomes, and lipids) in plasma. The assumption here is that the current flows radially into the space between the cell’s ventral surface and the substrate and then escapes between the cells. The current density is assumed to be consistent in the y direction, and the cells are disk-shaped objects with membrane surfaces and filled with a conducting electrolyte. Moreover, to identify the mechanisms and components involved at the cellular and molecular levels, we have started to investigate the plasma components (e.g., proteins, exosomes, and lipids) and individual cell types (e.g., T cells) separately. This investigation forms part of on-going studies that require further investigation before mechanisms may be suggested with a good degree of certainty."

I don't understand the biology, but it sounds like the anticipated mechanism could be much more than just interactions with the measurement probe.

 

But for 4000 wells that would be 0.0125 μL per well with an average of 2.5 cells. The smallest amount that a graduated pipette can discharge is about a micro litre. And that has to be discharged into a larger fluid volume otherwise it will just stick to one place by surface tension.

 

I sort of understand the biology but I never thought of blood cells as having a ventral (and presumably dorsal) side ??? I always thought of them as being without a top and bottom....it was a long time since I studied cell biology though (1980’s) perhaps things have changed? I haven’t read the paper yet...is it erythrocytes specifically they have used?

ETA: discovered an answer to one of my questions ...they remove red blood cells since this interferes with the signal ..gives the opposite result. Maybe this is due to lack of mitochondria or other organelles? Still bemused by ventral comment though.

 

I was amazed to read that the test is capturing healthy controls near an experimentally set "gray region" because these controls have relatives with Fibromyalgia (..!)

Although the results of the nanoneedle are striking , the next hurdle is to identify whether the test differentiates ME/CFS from other conditions. I do not know if the following is relevant however searching about generating "hyperosmotic stress" on PBMC cells (as stated in the paper) :

a) It appears that Hyperosmotic stress on PBMCs induces inflammatory cytokines :

https://www.sciencedirect.com/science/article/pii/S0014482797934765?via=ihub

b) On a paper named "The role of hyperosmotic stress in inflammation and disease" we read :

Link : https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3438915/

I think that this is interesting but also could be alarming in the sense that many types of disease could lead to the same result we have seen in the paper by Davis et. al ? In the paper linked above we find entries related to eye disease, diabetes, liver disease, inflammatory bowel disease and cardiovascular disease.

 

I certainly don't. Cells are not disk shaped.

The other thing I do not understand is why the electrodes are a few microns apart. Are they positive and negative, with current between, or is current transmitted to another electrode somewhere else? Cells are about 10 microns across so I would have thought current between two close electrodes would not pass through the cells much and certainly not between cells.

 

I think that paper just shows that you can find meaningless reviews on any idea you like these days. It seems to bear no relation to the reality of inflammatory disease whatever.

 

In which case I'm out of my depth so don't know.

 

Maybe this is where the capacitance come in. If the cells have a capacitive characteristic to them, then that may be what they mean when they speak of "membrane capacitance Cm". Capacitors have (virtually) infinite resistance (the real component of impedance), but finite reactance (the imaginary component of impedance). For capacitors, the reactance reduces the higher the frequency and the larger the capacitance. i.e. The obstruction to current is inversely proportional to f and C. I think many natural structures have capacitance characteristics. Maybe the cell membrane is behaving as a dialectric.

 

Yes I still remember that stuff from school but they do not seem to tell us about this.

And I am still puzzled as to why the pairs of electrodes are so close together.

 

According to figure 1 (A, B) The nano needle has two thin gold electrode layers sandwiched between three (Insulating?) oxide layers, presumably current flows between those two electrode layers via the fluid, but influenced by the close proximity of the cells, or even cell contact.

According to the scale on that figure 1, E), the nano needles themselves are spaced about 40 (Corrected from my earlier, higher figure) µm apart.

But perhaps I have misunderstood your point again

 

As I write this I see @Simon M has also commented on this. I presume you mean, by electrodes, the two gold layers within the sensor, separated by an oxide layer. The superimposed circuit diagram shows that the two gold layers and sandwiched oxide layer, comprise a capacitor, as well as having resistance within the gold layers, and presumably any connective wiring. So there is a CR circuit as the diagram shows. So an a.c. current will be able to pass between the gold layers, thanks to the capacitance; it will also be affected by the resistances ... impedance overall.

Anything coming into contact with this sensor tip, if it has its own resistance and capacitance, would effectively then become part of the circuit, giving modified circuit properties; the impedance would therefore change. If the fluid and its content presents higher resistance and/or lower capacitance, then presumably the impedance reading would be higher than for fluid with lower resistance and/or higher capacitance. (Impedance would be highest of all when the sensor is not immersed at all).

Edited for clarity.

 

Just a comment about lack of a complete description of the test equipment. This is typical of initial disclosures of technical advances with significant economic value. I've seen this with patents, trade secrets and non-disclosure agreements in a completely different context. Simply delaying the time until competitors can bring a challenge to market can be worth fortunes. Even non-profit organizations are affected by potential profits down the line which might indirectly benefit them or otherwise impact donors. Ron Davis has already signaled that there may be more than the 2 million sufferers in the U.S. NIH assumes, so we are talking about substantial value for equipment that can distinguish them. This will also have a major effect on insurance, and even debates about national healthcare.

When anything involves really large sums of money I warn people to "stand-by for maximum weirdness." When the numbers involve millions of people who are either fully or partially disabled the economic cost is staggering. We are getting into the range where it is impossible to distinguish rational economic self interest from psychosis, based on behavior alone.

(I've personally seen some extremes that would cross the line into criminal behavior, but have good reasons not to make them public due to laws about libel and slander. This is not simple cowardice, because I don't have important information and would be likely to make incorrect charges. Some of the information I do have may have been false and planted. Even good people will withhold information when faced with serious consequences, and there is no question bad people willing to distort perceptions do exist. )

I wish we could have rational discussions without the distortions introduced by narrow financial interests, some of them always hidden, but my view of current politics and economics suggests this is impossible. We will need patience and cunning to filter the information available until matters are firmly settled. We are piecing together a puzzle based on just a few pieces available to us. This is not the scientific ideal, but it is typical of reality.

 

Yep

 

It may be fairly typical of biomedical commercial development but it is not good science. And from my point of view if people are not prepared to publish clear good quality science I am inclined to ignore the stuff completely. I never concealed anything from anyone during my scientific career and I don't regret it.

 

This to me is one of the most interesting questions. I don't understand the ratio of plasma + salt soln to PBMC volume. Perhaps this is why the electrodes are close? Another reason could be that the closer together the electrodes, the higher the current, so background noise is much less. These are very tricky measurements and high impedance's are very difficult to measure in labs full of 60Hz background noise (paper describes needing faradic cage to minimise electrical lab noise).

Many people have done cellular impedance measurements on small circuit boards and add drugs to see the impedance change. These of course have larger geometries, but you can buy interdigitated circuit boards with 50um channels between the electrodes for ~$100 in one off quantities.

If one of the available types of commercial equipment could be adapted to incorporate the nanoneedle circuit, or the test adapted to use existing commercial equipment, then this test could be scaled up and replicated much quicker than just continuing with the nanoneedle alone. Indeed that is my hope. Just need $$$, ideas, and researcher availability + funding.

One experiment that would be really interesting would be to have Cornell measure cellular impedance of activated T-cells using commercial equipment (I read a paper on this) as they presented data at the NIH conference on activated T-Cells using Seahorse and also special staining. So you could tie the impedance change to two other measurables. The reason you activate T-Cells is to "stress" their energy complex. I guess it's probably not as simple as I write.........

But I guess the first job is to replicate the nanoneedle experiments in a larger group more diverse group using the existing setup before moving on to new experiments. Walk before you can run.

 

I don't expect Ron Davis to conceal matters for long, but he has already indicated there are aspects they don't understand. I'm sure you have been careful about making claims you were not ready to back up.

I'm also assuming we will see patent applications derived from this work before long. Once you start talking to university attorneys about intellectual property, things become more complicated, and they will forbid discussion of matters a scientist sees no harm in mentioning.

My background is not in commercial biomedical research, but I'm well aware of moral quandaries. I can understand all too well how Boeing got into the habit of downplaying the role of the MCAS features in the 737 Max 8, and how marketing types thought that making a few dollars by selling aircraft without the optional warning indicators about discrepancies from angle of attack sensors was good business. I'm sure they are reassessing that decision at present.

I left one job over disagreements concerning the Space Shuttle's thermal protection system, and regret that I could not do more to prevent the loss of Columbia years later. Perhaps, if I had not driven myself to the point of collapse, I might have made a difference.