**6. Lateral flow methods**

The development of lateral flow methods for detecting toxins has also led to the commercial availability of numerous kits for sensitive and rapid testing. Lateral flow methods employ capture antibodies that are "printed" on nitrocellulose membranes in a process akin to inkjet printing technology. Detection antibodies are labeled with visible materials, such as colloidal gold or colored latex beads. The sample is added to a reagent pad containing labeled toxinbinding detection antibodies and is wicked across the membrane. Toxin is retained by the capture antibody, which also concentrates the labeled detection antibody. A positive reaction is revealed as a colorimetric change and is presented as a line on the device. In general, lateral flow methods are qualitative and simply determine the presence or absence of neurotoxin. Sharma et al. [34] compared several commercial lateral flow devices for their capacities to detect toxin in food samples. They were able to detect BoNT/A and BoNT/B as low as 10 ng/mL and BoNT/E at 20 ng/mL in various liquids, such as milk, soft drinks, and fruit juices. Ching et al. [35] used the same mAbs described in the ELISA section above [24–26] in lateral flow devices to achieve sensitivities of 0.5 and 1 ng/mL for BoNT/A in buffer and milk, respectively. Although simple lateral flow tests have lower sensitivities compared to other methods, they produce rapid results and are most useful for the rapid screening of samples suspected of frequent contamination at relatively high level of BoNT. They have many applications and are ideal for field use by non-technical personnel. Self-contained and not necessarily requiring additional reagents or equipment, they can be easily interpreted in the field.

An innovative approach for toxin detection has recently been developed that combines antibodies with the amplification power of PCR, immuno-PCR (I-PCR) [36]. In I-PCR, template DNA is conjugated to the antibody, replacing a secondary antibody conjugated to the detection enzyme. Upon binding of toxin by the antibody, the presence of toxin is revealed using PCR. Chao et al. [36] described an I-PCR method for detection of BoNT/A with femtogram (10−15 g) sensitivity. These investigators compared competitive and sandwich ELISA to the I-PCR method. The I-PCR method was 103 –10<sup>5</sup> times more sensitive with LODs for the ELISA methods of about 50 fg. The use of I-PCR for highly sensitive detection of BoNT in food matrices or other biological and environmental backgrounds has yet to be reported (as of late 2015).
