Site-directed antibody immobilization techniques for immunosensors

• Published in: Biosensors & bioelectronics 2013-12, Vol.50, p.460-471
• Main Authors: Makaraviciute, Asta, Ramanaviciene, Almira
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Immobilized - chemistry; Antibody; Biological and medical sciences; Biosensing Techniques - methods; Biosensors; Biotechnology; Fundamental and applied biological sciences. Psychology; General aspects; GTP-Binding Proteins - chemistry; Humans; Immobilization techniques; Immunoassay - methods; Immunosensor; Methods. Procedures. Technologies; Site-directed immobilization; Staphylococcal Protein A - chemistry; Various methods and equipments
• ISSN: 0956-5663; 1873-4235
• DOI: 10.1016/j.bios.2013.06.060

Immunosensor sensitivity, regenerability, and stability directly depend on the type of antibodies used for the immunosensor design, quantity of immobilized molecules, remaining activity upon immobilization, and proper orientation on the sensing interface. Although sensor surfaces prepared with antibodies immobilized in a random manner yield satisfactory results, site-directed immobilization of the sensing molecules significantly improves the immunosensor sensitivity, especially when planar supports are employed. This review focuses on the three most conventional site-directed antibody immobilization techniques used in immunosensor design. One strategy of immobilizing antibodies on the sensor surface is via affinity interactions with a pre-formed layer of the Fc binding proteins, e.g., protein A, protein G, Fc region specific antibodies or various recombinant proteins. Another immobilization strategy is based on the use of chemically or genetically engineered antibody fragments that can be attached to the sensor surface covered in gold or self-assembled monolayer via the sulfhydryl groups present in the hinge region. The third most common strategy is antibody immobilization via an oxidized oligosaccharide moiety present in the Fc region of the antibody. The principles, advantages, applications, and arising problems of these most often applied immobilization techniques are reviewed. •Antibody orientation on the surface is a pivotal step in immunosensor development.•Three most common antibody site-directed immobilization strategies are reviewed.•Immobilization via Fc binding proteins, oxidized oligosaccharides and sulfhydryl groups of the hinge region.•The principles, advantages, and arising problems of these techniques are discussed.•Comparison of different antibody immobilization strategies is presented.