Three Dimensional Substrates

Polyacrylamide-coated slides form a three-dimensional substrate for protein microarrays. Much like the polyacrylamide gels used for electrophoresis, they are reported to provide high probe loading capacity and a hydrophilic, solution phase-like environment, which is thought to preserve protein probe function [3-6]. Consequently, this flexible and versatile platform is well suited for many types of proteins and has been used to develop a variety of assays in microarray format. A three-dimensional hydrogel coating preserves the native three-dimensional structure of proteins, thereby maintaining stability and functionality. The reactive chemistry is stable and remains active even during very long spotting (also know as "printing") runs. Agarose is an economical, protein-friendly medium that reduces the background of the slide significantly, which can interfere with signal detection of low-abundance proteins or pep-

tides. There are other proprietary three-dimensional porous polymer surfaces, such as those produced by Full Moon Biosystems, that claim to provide a high level of sensitivity and specificity for fluorescence-based assays. All seem to have similar properties, which include covalent multifunctional bonding sites that provide high binding specificity by covalent attachment to surface functional groups.

Nitrocellulose is the best-characterized protein-binding chemistry that has been adapted successfully to microarray fabrication. Nitrocellulose membranes are used for immobilizing a wide variety of molecules, such as nucleic acids in Southern and Northern blots, as well as proteins in Western blots. Nitrocellulose makes an ideal surface for a wide variety of assays, including antibody-based assays, due to its nonspecific affinity for proteins. Nitrocellulose is usually coated onto a nylon backbone to produce membranes. It has also been coated on activated glass microscope slide, and as such has become a very popular tool for the fabrication of protein microarrays. A variety of nitrocellulose-coated slides are currently available which are ideal for microar-rays because of their high binding capacity per unit volume. At one end of the spectrum, the nitrocellulose surface may be as thin as 5 |im (Path Slides, GenTel), and at the other, the membrane can be as thick as 150 |im (SuperNitro, Telechem). In the case of nitrocellulose-coated slides, it is critically important to pick the correct thickness for the application. The thickness of the coating chosen will be influenced greatly by the type of microarray printing that is to be used.

Project Management Made Easy

Project Management Made Easy

What you need to know about… Project Management Made Easy! Project management consists of more than just a large building project and can encompass small projects as well. No matter what the size of your project, you need to have some sort of project management. How you manage your project has everything to do with its outcome.

Get My Free Ebook


Post a comment