Biosensors for the Detection of Waterborne Pathogens - Waterborne Pathogens: Detection Methods and Applications - page 191

is one of the most critical steps in biosensor development because biosensor
performance (sensitivity, dynamic range, reproducibility, and response time)
depends upon the recognition element retaining its original properties sub-
sequent to immobilization. For example, some methods of antibody immo-
bilization are nondirectional, and therefore some of these antibodies may thus
end up immobilized in orientations such that target recognition is difficult.
Stability of the immobilization as well as the extent of surface coverage are
also important factors to consider in the selection of a strategy.
Some examples of existing immobilization strategies include biomolecule
physisorbtion, entrapment and encapsulation into polymers or membranes,
silanization, and self-assembled monolayer (SAM) formation coupled to
biomolecule cross-binding or covalent bonding using linkages such as pro-
tein A or protein G, biotin-streptavidin or N -hydroxysulfosuccinimide and
1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride. Details of
these methods can be found in the excellent topic Bioconjugate Techniques 19 as
well as in other recent reviews. 20,21 In addition, the modified surface has to
be inert and biocompatible so that it does not affect the sample composition
or integrity in any way; it should also guarantee a constant signal baseline.
Blocking of unmodified surface is also required, using, for example, bovine
serum albumin (BSA) to prevent nonspecific binding.
The easiest way to immobilize antibodies is to unspecifically deposit or
adsorb them on the sensing surface. Nonetheless, most authors maintain
that directed immobilization by chemical conjugation or crosslinking of
precise functional groups in the antibody (i.e. amines, carboxylates, carbo-
hydrates) better preserves their integrity and functionality, while promoting
more organized structures. This was found, for example, in a recent study of
the best immobilization strategies for Cryptosporidium . 22
7.3. TRANSDUCTION METHODS
7.3.1. Optical
Many optical biosensors rely on fluorescence, which was covered in Chap-
ter 5. Lateral flow assays (LFAs) are a common type of sensor, well-known
for the home pregnancy test example application. In this type of assay the
test sample flows along a solid substrate past a functionalized area that would
capture the target of interest, leading to a color change. Most examples of
the above types of detection approach were covered in Chapter 5. This
chapter concentrates on those methods in which the binding of the analyte