Aptamer-antibody sandwich lateral flow test for rapid visual detection of tetrodotoxin in pufferfish

Abstract

Tetrodotoxin (TTX) is a highly potent marine toxin which can cause severe poisoning following consumption of contaminated fish and seafood. Thus, a sensitive, reliable and simple test is required for rapid screening of samples and prevention of intoxication. Herein, we translated a previously reported microtiter plate hybrid aptamer-antibody assay into a rapid lateral flow assay (LFA) test. The test relies on an aptamer immobilized on the membrane and an antibody conjugated with gold nanoparticles to provide a visual result when TTX is present in the sample. The optimized test is simple (one-step), rapid (<20 min), highly sensitive (visual limit of detection of 0.3 ng/mL TTX in buffer corresponding to 0.78 mg TTX/kg tissue), specific, reproducible and with long storage life. It was validated by analyzing contaminated pufferfish tissue extracts and it successfully detected TTX below the current limits set by official bodies. The analysis performed with this device in combination with a simple LFA reader for quantification was in excellent agreement with other established methods, further demonstrating the value of this test as a simple, low-cost and reliable analytical tool to ensure food safety, protect human health, and broaden the knowledge on the correlation between biological parameters and environmental data.

Keywords: Antibody-aptamer sandwich; Aptamer; Gold nanoparticles; Lateral flow assay; Pufferfish; Tetrodotoxin.