Antibodies provide a remarkably specific way to recognize invading pathogens such as bacteria, viruses, or tumor cells. It is no wonder that antibodies are widely used as critical reagents in medical and biological research laboratories----they provide exquisitely specific and powerful ways to detect specific molecules in complex mixtures, such as bodily fluids. Antibodies are also widely used to separate and purify specific proteins of biomedical interest.

Current ways of using antibodies for detection and purification rely on linking methods that greatly reduce their ability of antibodies to bind to their specific targets, which are known as antigens.

In nature, however, antibodies, which are Y-shaped molecules, are properly oriented and tethered by their constant (Fc) regions (located at the end of one leg of the Y----see Figure 1), in a way that maximizes their ability to bind to specific antigens.

Now, in a recent study to be published in Biotechnology and Bioengineering, a team of researchers led by Hsuan-Chen Wu's work and UMBI's Dr. William Bentley has described a more natural way to orient and assemble antibodies----a entirely biologically based, bottom-up method of fabrication which can correctly orient one or more different types of antibodies at specific locations on a variety of devices.

Figure 1. Schematic of the biofabrication method for antibodies.. The constant region of antibodies are naturally bound to modified protein G molecules which are enzymatically coupled to chitosan. This orients the antibodies in a natural way, which optimizes binding to their specific antigens. The chitosan-protein G-antibody assembly can be deposited at specific desired locations. In this way, multiple antibodies can be linked to devices, including ultra-miniature nanoscale devices.

Multi-functional lab on a chip devices----which can detect several different antigens at once----are among the most exciting applications made possible by the new method. This is made possible by using chitosan, which can be deposited at specific chip by electrical means.

Dr. Bentley holds a joint appointment as a professor in UMBI's Center for Biosystems Research (CBR) and as Chair of the University of Maryland's Fischell Department of Engineering at College Park. Other CBR and Fischell authors on the paper include first author Hsuan-Chen Wu, Xiao-Wen Shi, Chen-Yu Tsao, Angela T. Lewandowski, Rohan Fernandes, Chi-Wei Hung, and Gregory F. Payne, who is Director of CBR. Dr. Payne was instrumental in the development of chitosan, a key material used in the new assembly process. The paper is also co-authored by Philip Deshong of the University of Maryland, Eiry Kobatake of the Tokyo Institute of Technology, and James J. Valdes of the U.S. Army Edgewood Chemical Biology Center.

The new assembly method relies on the special properties of chitosan, a converted waste material derived from the shells of crabs or shrimp. Chitosan can be deposited on solid surfaces at specific locations by either changing the pH of the solution, or by means of an electrical current, and serves as a universal scaffold that can bind a variety of biological materials, including proteins, to those surfaces.

The new assembly uses a specially engineered derivative of protein G, a biological material from bacteria that spontaneously binds tightly to the constant region of antibodies. The engineered protein G has a special site that is linked to chitosan by means of a specific enzyme. The chitosan-protein G-antibody assembly can then be deposited at specific locations on various types of solid surfaces.

The study demonstrated that functional antibodies can be deposited on three types of devices, including electrode addresses of microfluidic devices, multi-well plastic plates, and woven materials. The antibody assemblies prepared in this manner were shown to function quite effectively in binding their specific antigens. Antibodies that were air dried remained 95% functional after 7 days, indicating that devices with the antibody assembly can be prepared and stored for later use.

Many applications are envisioned for the new devices, including the fabrication of ultra-miniaturized, multi-function detectors in nanoscale devices.