Rapid visual tests: fast and reliable detection of ochratoxin A

Abstract

This paper reviews the early detection strategies that have been employed for the rapid monitoring of ochratoxin A (OTA) contamination of food. OTA, a mycotoxin mainly produced by some Aspergillus and Penicillium species, is found in cereals, coffee, wine, pork and grapes. To minimize the entry of this mycotoxin into the food chain, rapid diagnostic tools are required. To this end, the potential use of lateral flow devices has also been developed. In this study, we analyze the robustness of test strips using published methods for colorimetric detection. Different test formats are discussed, and challenges in the development of lateral flow devices for on-site determination of OTA, with requirements such as robustness, speed, and cost-effectiveness, are discussed.

Keywords: clean-up immunoassay; colorimetric test; flow-through test; lateral flow test; ochratoxin A.

Figure 1

Competitive assay in test strip format (free-standing test strip). (A) The principle of the method, modified from [11]. The toxin recognition sites of the specific antibody conjugated to a detection system (usually colloidal gold) located in the conjugated pad will be blocked by the toxin present in the sample, thus preventing the antibody from being fixed on the test line. In the control line, an anti-antibody (usually goat anti-mouse antibody) will retain charged and uncharged conjugated antibodies, thus providing the positive control. The intensity of the test line will be inversely correlated to the toxin concentration in the sample. (B) Schematic diagram of simple and multiple detection strips. The two test lines contain different toxins conjugated to the membrane-bound protein. The conjugate pad contains specific antibodies to each of the corresponding toxins.

Figure 2

Principle of competitive immunoassay with conjugated toxin, modified from [24]. (A) The membrane is coated with the first antibody (usually a goat anti-mouse); (B) The second antibody (usually a monoclonal anti-toxin) is then fixed; (C) The membrane is then placed in contact with the sample. If the sample contains the specific toxin, the toxin links to the specific antibody; (D) A detection element conjugated with the toxin (usually HRP) is then added to the membrane. The amount of conjugated toxin that can be fixed is inversely correlated with the amounts of toxin present in the sample; (E) The non-fixed conjugated toxin is rinsed away before adding a developing product (F).

Figure 3

The principle of the simultaneous immunoassay method for aflatoxin B1 (AFB1) and OTA in a flow-through assay. Two types of reagent spots are prepared, one containing the conjugates for AFB1, the other for OTA. The detection process is the same for both samples.

Figure 4

Set-up of the flow-through column connected with a clean-up column.