Location: Foodborne Toxin Detection and Prevention Research
Project Number: 2030-42000-053-014-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 15, 2025
End Date: Oct 31, 2028
Objective:
Foodborne outbreaks from pathogens and their toxins are a tremendous burden on the US economy with an estimated cost of $75 billion dollars impacting U.S. farmers, producers, and consumers. Bacterial toxins are among the most notorious agents responsible for foodborne illnesses. They are produced by bacteria and can contaminate food at various points from production to consumption. The health impact of these toxins ranges from mild gastrointestinal discomfort to fatal conditions. The most well-known examples include botulinum toxins, Staphylococcus enterotoxins, and Shiga toxins. Identifying the contaminated sources early is the key step in mitigating their effects. Existing diagnostic technologies, such as animal-based cytotoxicity assay, enzyme-linked immunosorbent assay (ELISA) and mass spectrometry analysis either lack the needed sensitivity or require expensive infrastructure, which is not applicable for use in the field and resource-limited settings.
The objective of this cooperative research project is to develop ultrasensitive (< 1 pg/mL), low cost (< $3/test), field-deployable, and one-step diagnostic sensors for rapid (< 30 min) detection of foodborne toxins in food and environmental samples without sophisticated laboratory infrastructure. The novel and critical detection platforms developed may be further applied to the detection against a variety of high-impact pathogens, toxins, and potentially mycotoxins that can contaminate high-value agricultural commodities which will have direct impact in protecting U.S. agriculture, farmers, producers, consumers and ensuring economic prosperity.
Approach:
The proposed project will use reagents including toxins and antibodies/nanobodies developed by ARS scientists to build antibody/nanobody-functionalized nanoparticles for rapid electronic detection (Nano2RED) of foodborne toxins. In Nano2RED, gold nanoparticles (AuNPs) will be covalently coated with high-density ligands, such as streptavidin proteins, and then functionalized with target-specific binders, such as monoclonal antibodies (mAbs) or nanobodies through a biotin-streptavidin reaction. Acting as multivalent, high-affinity, in-solution sensors, the AuNPs actively bind target protein markers upon mixing and form clusters that subsequently precipitate from the solution. The signal transduction of target protein molecule recognition involves multiple chemical and physical steps including 1) centrifugation (~5 min) immediately after the mixing of protein molecule samples, thus both significantly improving the reagent concentration at the tube bottom and accelerating assay signal transduction; 2) room temperature incubation for 5 to 20 min; and finally, 3) a brief vortex agitation (5 sec) to return the monomer AuNPs back to the solution without breaking the AuNP clusters “glued” together by the target proteins, thereby ensuring specificity. The target protein concentration dependent modulation of AuNP clustering allows accurate signal readout by probing the optical extinction of free-floating AuNPs. The color change can be precisely quantified using a portable electronic detecting system (PED) comprising a light-emitting diode (LED), photodiode, a battery and supporting circuit. The PED readout signals are calculated in reference to positive control (PC) and negative control (NC) samples to obtain a normalized electronic signal for each protein concentration. After assessing assay sensitivity and specificity using
relevant food matrices, field deployable systems will be set up for on site testing.