Food Biochemistry and Food Processing

31 Emerging Bacterial Foodborne Pathogens and Methods of Detection 733

The plate is blocked and washed, and then an anti-
body (primary antibody) specific for the pathogen is
added to the well. After a suitable incubation period,
the plate is washed to remove unbound antibodies. A
secondary antibody conjugated to an enzyme that
converts a colorless substrate to a visible product is
then added. The secondary antibody has specificity
for the primary antibody, so if the primary antibody
is bound to any antigen, the secondary antibody will

bind to the primary antibody. Finally, the plate is

washed again to remove any excess secondary anti-

body, and a substrate is added that turns color in the

presence of the enzyme. The intensity of the color is

proportional to the amount of pathogen present in

the original sample (Hess et al. 1998). In one varia-

tion of this assay, the plate can first be coated with

an antibody specific for the pathogen. This allows

the pathogen in the sample to be concentrated on the

Figure 31.4. A.Conventional IgG antibody and various possible recombinant fragments derived from the antibody.
Fab is composed of the first constant domain and the variable domains of both the heavy and light chains. scFv is
composed of the variable light and heavy domains connected and stabilized by a flexible peptide linker. VH is derived
from only the heavy-chain variable region. B.Heavy-chain antibody (HCAb) and its possible recombinant derivatives.
The heavy-chain antibody is found only in the camelid family. VHH is derived from the variable region of the HCAb.