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Zeptosens Michael Pawlak on High Throughput Analysis of Cell Signaling

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At A Glance

Name: Michael Pawlak

Age: 42

Position: Director of Protein Microarrays, Zeptosens, Switzerland

Background: Investigator, Bioanalytical Research Department, Ciba Pharma, later merged into Novartis Pharma, 1994 to 1999.

Postdoc, Swiss Federal Institute of Technology, Lausanne, Switzerland, 1991-1994

PhD in biophysical chemistry, Biocenter, University in Basel, Switzerland, 1988-1991.

 

How did you get into studying cell signaling and high-throughput protein profiling?

During my education, I was more related to classical high-throughput screening but not on the signaling profiling screening. But the main point was that we had contact to a lot of customers from pharma which actually had this need and couldn’t do high-throughput profiling of downstream signaling with classical or conventional methods like western blots. They were actually looking for technologies. And so we had the technology to do that so we met and then came to this kind of solution which is very adapted to the classical high-throughput screening in pharma — not in the field of signaling, but in receptor-ligand interaction.

What did you do before joining Zeptosens?

I did my PhD in Basel in Switzerland in the biocenter of the university. This was in biophysical chemistry, so this was related to protein-protein and protein-membrane interactions. From there I went as a postdoc to the Federal Institute of Technology. There I was working on setting up biosensing methodologies, looking for protein interaction, making assay developments, not the classical microarrays, but kind of surface plasma resonance methods and electrical methods combined into new sensing schemes. Biosensing was a chip-based analytical test. You had pharma-relevant receptor molecules on this chip surface you probed for small ligand interaction on the surface. This was in developing methods to screen for new drugs.

Where did you go from there?

Then I went to Ciba Geigy in Bartel, Switzerland, which was a pharma company that has now merged into Novartis. I entered their bioanalytical department in 1994. We developed there this optical method which we apply now in our spinoff company which is the protein microarray readouts technology. The optical method is a glass chip which is excited on the surface optically, and you can bring protein microarrays on the chip and do a fluorescent readout on these microarrays. It’s a method which, compared to conventional technologies, is very sensitive and only reads on the surface, not in solution.

Later, we did the spinoff out of Novartis to the company I’m with now, which is Zeptosens. We developed this microarray technology which we can use for proteomics for this signaling profiling in high-throughput.

What does the microarray technology involve?

We use a special form of protein chip — it’s an array of samples instead of an array of antibodies. We call it a lystate array — it’s an array of lysed biological samples. And you can probe these arrays with antibodies for specific cell signaling molecules. We sell these lysate arrays. It’s different from SELDI technology — SELDI is a mass spec technique which relies on detection of mass. We use this fluorescent readout which is much more sensitive. It’s much more applicable for high throughput. We can put hundreds of samples onto a small array. For SELDI, it’s only a few.

How is the fluorescence read?

We have developed our own reader which reads the fluorescence emitting from this chip. It’s an Imager. It’s very new. Not only the system itself, but the whole approach is rather new — the chip with these arrays and the readout, and the protocols that we develop for it. We sell this and it’s applied in the pharma industry, in proteomics environments.

How do you go about analyzing the data that you get from this?

There’s an automated readout. You put in a number of chips into the machine, and it reads out all the stuff automatically, and then you have to do a computer analysis on this data.

We have also our own software for this. It’s called Zeptoview Pro. Our reader is called the Zepto Reader, because we detect zeptomoles, which indicates very small amounts of molecules. A zeptomole is 10 to the minus 21 moles, which is only 600 molecules on a microspot. I would say it’s the most sensitive system. The samples which you put on a chip have all the proteome on there, and you can really detect a low abundance molecule out of this mass of other matrix proteins which are present.

How is this technology applied?

It can be applied in early drug discovery, for instance, looking for drug candidates which are blind to cell systems. You can then study the down cell signaling effects on not only one but many protein markers in parallel. For cell signaling, what you measure is actually the expression of downstream signaling markers in a cell in this live sample which is deposited as a spot on the chip. So, by addressing different sorts of antibodies to the array, you can then generate signals from different downstream signaling markers, and you do that in parallel. So you measure in parallel hundreds of samples and you measure a large number of different proteins all at once. Pharma people use this technique now for screening, for early selection of candidates.

Do you see this technology as competing with other chip technologies that use mass spectrometry?

It’s complementary to it. What it is most comparable to is the conventional Western blot which is years old, very cumbersome and time consuming. This is a method that has the benefit of very rapidly analyzing a lot of samples and analytes in a short time. So for instance, to analyze 500 patient samples or discovery samples with 50 different analytes would take two weeks with this method, compared to at least half a year of time with Western blots.

What do you see as the main advantages of this method over mass spec methods?

It’s a fast method, it’s easy to use, it’s fully automated. I wouldn’t say it’s replacing mass spec, it’s just a complementary thing. Because what we do is profiling in very complex mixtures, and do it very quantitatively. And this is not possible with such high sensitivity with mass spec. So certainly, a higher sensitivity, a better precision and the option for quantification. You can measure the level of cell signaling proteins relative to a disease. We can compare a diseased sample, such as a breast cancer tissue with a normal tissue. You can detect the differences in levels of these proteins, and we do that very accurately and sensitively. If a certain protein is only present at a very low level, we can address it and also detect changes. Mass spec has its limits.

What are you working on now?

We study signaling pathways in customer studies, in a kind of service. For example, we study apoptosis. What we do is actually confirm hypotheses on certain systems. A hypothesis might be that a disease follows a certain pathway. We have done studies of UV damage, DNA damage of certain cell lines. Classical methods only measured one protein and we confirmed that a number of proteins aligned in this pathway showed the expected activation. We enter now in this kind of proteomics application to an area where we do not look at just one protein but at a whole set of proteins which was selected with a certain theoretical background from biology. With the conventional methods, the researchers can only test an individual protein because they don’t have the time and the budget to make the measurements for all of the proteins with the samples and often they don’t have the reagents for it. We can now very rapidly investigate whole cascades of the many proteins involved in these hypotheses and actually prove these hypotheses. We’re testing, cascades, pathways, and also interactions of proteins in different pathways. There’s now the idea that a protein from one pathway can have an interaction with a protein from another pathway. There’s really not published proof of this, but we can now confirm this hypothesis. At the end, it could enter into a kind of global analysis. In the end, in principle, you could investigate not tens, not hundreds but thousands of proteins if you have all the antibodies for them.

What are you looking to do in the future?

Spread the technology. We’ve gone to some trade shows, and we have people in direct contact with the pharmaceutical companies.

 

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