Recombinant antibodies and their use in biosensors

Abstract

Inexpensive, noninvasive immunoassays can be used to quickly detect disease in humans. Immunoassay sensitivity and specificity are decidedly dependent upon high-affinity, antigen-specific antibodies. Antibodies are produced biologically. As such, antibody quality and suitability for use in immunoassays cannot be readily determined or controlled by human intervention. However, the process through which high-quality antibodies can be obtained has been shortened and streamlined by use of genetic engineering and recombinant antibody techniques. Antibodies that traditionally take several months or more to produce when animals are used can now be developed in a few weeks as recombinant antibodies produced in bacteria, yeast, or other cell types. Typically most immunoassays use two or more antibodies or antibody fragments to detect antigens that are indicators of disease. However, a label-free biosensor, for example, a quartz-crystal microbalance (QCM) needs one antibody only. As such, the cost and time needed to design and develop an immunoassay can be substantially reduced if recombinant antibodies and biosensors are used rather than traditional antibody and assay (e.g. enzyme-linked immunosorbant assay, ELISA) methods. Unlike traditional antibodies, recombinant antibodies can be genetically engineered to self-assemble on biosensor surfaces, at high density, and correctly oriented to enhance antigen-binding activity and to increase assay sensitivity, specificity, and stability. Additionally, biosensor surface chemistry and physical and electronic properties can be modified to further increase immunoassay performance above and beyond that obtained by use of traditional methods. This review describes some of the techniques investigators have used to develop highly specific and sensitive, recombinant antibody-based biosensors for detection of antigens in simple or complex biological samples.

Antibody Fragments and Their Purification by Protein L Affinity Chromatography

“…Ab-fragments are especially amenable to genetic manipulations (see below) or covalent modifications (e.g., [13,30]) aimed at improving their affinity, stability, and function. Antibody fragments have been linked with enzymes [31], toxins [32], and radionuclides for cancer treatment [33][34][35], as well as viruses for gene therapy, liposomes or nanoparticles for improved drug delivery, and dye or other sensing substances [11,[36][37][38][39]. Figure 2 depicts some Ab-fragments.…”

The development and optimisation of nanobody based electrochemical immunosensors for IgG

“…Antibodies may also bring issues of cross reactivity which cannot be identified until after production. These technical shortcomings are in addition to the ethical issues raised by their reliance on animal use and the financial issues raised with their associated cost [4,5].…”

De novoisolation of antibodies with pH-dependent binding properties

“…6 These techniques take advantage of the high specificity of the interaction between a mAb and its target to allow the detection of a defined molecule even in complex biological samples. 7 The specificity of mAbs has, for instance, been exploited to develop kits allowing a rapid identification of pathogenic viruses or bacteria for human and cattle diseases. 8 In addition, antibodies are powerful purification tools and antibody-coupled resins are used to purify not only single proteins, but also enzymes, drugs, proteins complexes and lipids by immunoaffinity chromatography.…”