Biorecognition

The biorecognition elements are biologics such as enzymes, antibodies, and even biological cells and microorganisms that selectively recognize an analyte. They are often immobilized to increase their local concentration near an optical sensing element and to allow them to be reused. Some of the molecular bioreceptors used for biorecognition in biosensitizing are described here.

Enzymes. The use of an enzyme as a biorecognition element utilizes its selectivity to bind with a specific reactant (substrate) and catalyze its conversion to a product. This enzyme-substrate-catalyzed reaction, also discussed in Chapter 3, is often represented as

In addition to providing selectivity, the reaction of certain analytes/substrates with enzymes can also provide optical transduction by producing a product that absorbs at a different wavelength (change in absorption), or is fluorescent (fluorescence sensor). Alternatively, the product of the enzyme-catalyzed reaction can interact with a dye (an optical sensing element such as a fluorescence marker) to produce an optical response.

Antibodies. Antibodies, as discussed in Chapter 3, are proteins that selectively bind with an antigen or hapten (analyte) because of their geometric (site) compatibility. Very often an antibody-antigen pair's selective association in terms of their conformational compatibility is represented as a lock (antibody) and key (antigen) combination, as shown in Figure 9.2. This specific physical association can also produce an optical response that can be intrinsic such as a change in the optical property of the antibody or the antigen

ANTIBODY SPECIFICITY heavy chain

antigen light chain

ANTIBODY SPECIFICITY heavy chain

antigen light chain

An individual animal can make billions of different antibody molecules, each with a distinct antigen-binding site. Each antibody recognizes its antigen with great specificity.

Figure 9.2. Schematic representation of antibody-antigen selective recognition. (Reproduced with permission from http://www.accessexcellence.org/AB/GG/antibodies.html)

as a result of association. Alternatively, an optical transducer (such as a fluorescent marker) can be used to tag the antibody or the antigen.

Lectins. Lectins are proteins that bind to oligosaccharides or single-sugar residues as well as to some glycoproteins such as immunoglobulins. Therefore, the lectins can act as biorecognition elements for these analytes. For example, concanavalin A in its A-form has been extensively used for its specific binding with a-D-mannose and a-D-glucose residues in a glucose sensor. In a glucose sensor utilizing concanavalin A, the lectin is immobilized on a sepharose film coated on the interior walls of a hollow fiber (Schultz et al., 1982; Boisde and Harmer, 1996). Furthermore, it is liganded (conjugated) to dextran labeled with a fluorochrome, fluorescein-isothiocyanate (FITC). The glucose as an analyte diffusing through the hollow fiber displaces dextran from concanavalin A. The fluorescently labeled dextran then migrates to the area illuminated by light, being conducted through an inner solid optical fiber, to produce detectable fluorescence.

Neuroreceptors. These are neurologically active compounds such as insulin, other hormones and neurotransmitters that act as messengers via ligand interaction. They are also labeled with a fluorescent tag to produce an optical response through chemical transduction.

DNA/PNA. The specificity or complementary base pairing (that provides the basis for the DNA double-helical structure) can be exploited for recognition of base sequence in DNA and RNA (Kleinjung et al., 1998). An example is a DNA microarray (detailed coverage in Chapter 10) that consists of micropatterns of single-stranded DNA or finite-size oligonucleotides immobilized on a plate. They act as biorecognition elements by forming hydrogen bonds with a specific single-stranded DNA or RNA having a complementary base sequence. This process of base-pairing to form a double-stranded DNA is called hybridization (see 8.5.4). Another example of biosensing utilizing the hybridization in DNA is provided by a molecular beacon sensor, discussed below.

Recently, remarkable sequence specificity has been reported using peptide nucleic acids (PNAs) as biorecognition elements (Wang, 1998; Hyrup and Nielsen, 1996). The PNAs provide the advantage of a neutral backbone and correct interbase spacing to ensure that the PNAs bind to their complementary sequence with higher affinities and with specificity comparable to oligonucleotides.

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