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Carbohydrate Chips Take a Step Forward with Help from ISU s Nicola Pohl

Nicola Pohl
Assistant Professor, Chemistry
Iowa State University

At a Glance

Name: Nicola Pohl

Title: Assistant Professor, Chemistry, Iowa State University

Professional Background: 2000 — present, assistant professor, Department of Chemistry, Iowa State University;

Education: 1997 — PhD, chemistry, University of Wisconsin, Madison; 1991 -BA, English and Religion, Harvard College.

Awards: 2005 — Alfred P. Sloan Research Fellow; 2004 — National Science Foundation CAREER award; 2003 — Research Corporation Cottrell Scholar

A paper entitled "Fluorous-based carbohydrate microarrays" published this week in the Journal of the American Chemistry Society shows that although DNA and proteins remain the biomolecules of choice for arrays these days, scientists are working hard at making carbohydrate microarrays accessible and easy to use.

Carbohydrate microarrays are of themselves nothing new. An Israel-based company called Glycominds has already managed to commercialize a carbohydrate chip. But according to Nicola Pohl, a chemist at Iowa State University and the lead author of the JACS paper, traditional microarray slide chemistries have limited the utility of carbohydrate chips.

Rather than build on the current methods of binding carbohydrates to surfaces, Pohl and her associates decided to throw the old method out, and focus on creating a method based on non-covalent fluorous-based interactions.

In addition to designing the resulting slide chemistry, Pohl is now busy at work readying it for commercialization with Fluorous Technologies, a Pittsburgh-based chemical technology company. To learn more about the new carbohydrate-friendly chemistry and its future, BioArray News spoke with Pohl this week.

Where have you been the past few years and how did you get started on this research?

I started out at Iowa State as an assistant professor in August 2000 and I was interested in carbohydrates. The problem there is we don't know as much about carbohydrates and proteins in large part because we don't think we have the same sort of tools available to study carbohydrates that have become available in the last 20-30 years to study proteins and DNA. And the fact that there are carbohydrate microarrays that just started coming out — technically since 2000 — is indicative of that, for instance. Part of it was a drive to be able to make carbohydrates more easily and then also, because they are so hard to get synthetically, to array them very easily. One thing in particular that we are interested in is glucosaminoglycans.

We don't have any arrays [for glucosaminoglycans] and one of the problems for trying to make arrays like that is that the standard method uses covalent chemistry. The standard way of making slides for DNA, for example, is to covalently link the DNA to the actual glass slide. That means you need some sort of unique group on the DNA to do that chemistry. The problem is that that unique group on the DNA can sometimes be a naturally occurring group on a carbohydrate and that complicates the chemistry.

If you were just to array carbohydrates on a standard surface, what would happen?

Probably they would just come right off again. So what we've done is, instead of using this covalent chemistry, we've developed a Teflon-like tail on our sugars. And the neat thing about this Teflon-like tail is that you can not only use it to help purify the compounds when you make them, but you can also use that same tail to directly array it on a glass slide that's been coated with something that makes it Teflon-like. So, literally, water beads up on the surface like a Teflon pan. And so that's how the surface is different from conventional surfaces or slides. It's been coated with something that makes it highly fluorinated, fluorocarbons. Normally, proteins and buffers and everything like that won't stick to the surface but our sugars have this tail that will allow them to stick to the surface. And so wherever we spot the sugar on to this Teflon-coated slide, essentially the sugar will stick.

How did you discover the surface chemistry?

The driving force was actually trying to find a quick way of making the carbohydrates themselves. DNA is usually made on solid phases. And we tried the solid-phase chemistry with carbohydrates and never had much luck with it. So then we were looking for another way to make the carbohydrates with the ease of purification that solid phase would have, and that's how we landed on these sort of fluorous tags that were developed in the mid-1980s.

We thought that if we developed a new method that would allow us to use the tag directly, it would save us a lot of steps, which is especially [useful] if you are trying to make small libraries of compounds to array.

Why is there a need for this specific technology and for carbohydrate arrays in general?

There have been some other carbohydrate arrays using the same sort of techniques that are used to make DNA and protein arrays. The drawback for those is that many of the naturally occurring carbohydrates have groups on them that make the attachment chemistry very difficult — so it's time consuming. Because we're not actually doing any bond-forming reactions on the surface, it's a lot simpler to be able to make our arrays than what's out there currently. And we literally just spot the slide, we don't need to incubate, we don't need to figure out optimal pH and time and all that other stuff because we are not carrying out a covalent bond reaction.

Generally, [this technology] has been used to screen for antibodies, because carbohydrates are found on the surface of human cells, pathogens, viruses. And a lot of our immune responses against those viruses and pathogens are mediated by carbohydrates. So people are interested in carbohydrate arrays to be able to screen to see what kind of carbohydrates we generate an immune response against. For example, somebody who's been exposed to a [Staphylococcus] aureus infection — what kind of antibodies do they generate against that that bind the carbohydrates? You can take that patient's antibody sample and see which ones bind, and then it gives you a clue to what kind of vaccine you might want to develop, based on that carbohydrate structure.

People are also just trying to figure out the basic biochemistry of immunology. Since so many of our immune system proteins also bind to carbohydrates, we don't actually know which carbohydrates many of them bind to. And so there's the basic science aspect of figuring out what is the specificity of all these kinds of carbohydrate-binding proteins involved in the immune system in neuron communication, for example.

Where are you going to go with this?

Now what we'd like to do is expand the scope of it because we've demonstrated it with small carbohydrates. We are looking at glucosaminoglycan structures and we also want to use that same fluorous tag to start making small libraries of carbohydrates that are implicated in immune responses. We are in the process now of making small libraries of carbohydrates that we can quickly make slide arrays to be able to screen against various immune system proteins.

What kind of equipment are you using for this?

We are using standard DNA microarray equipment. So anyone that has access to a DNA microarray facility could use it. We are literally using our DNA microarray facility across the street.

So would you be willing to provide these slides to researchers?

We are in negotiations with Fluorous Technologies right now, for them to start selling these slides. We have a National Institutes of Health grant that just got funded to try and commercialize this technology. [According to an NIH database, the $170,272 Phase I STTR, "Fluorous Solution-Phase Synthesis of Peptides and Oligosaccharides," runs through July 2006 — Ed.]

When do you anticipate a product could be ready?

[Fluorous Technologies] are hoping to have beta-test versions for customers by the end of the year actually — so in a couple months. Part of that is to be able to do this at a large scale and get consistency of process.

What kind of features will be arrayed on these chips?

There are way more carbohydrates than there are possible DNA sequences. So there's no way you could have one chip that would have every possible carbohydrate on it. It tends to be much more problem specific. It's really application specific. And since this paper has come out I've heard through the grapevine that a lot of people in the pharmaceutical industry have been using these tags to facilitate their execution. They are really excited about this because now they've got a method where they can directly array and screen the compounds in their bioactivity assays.

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