The discovery of in-vitro systematic evolution of ligands by exponential enrichment (SELEX) process has
considerably broaden the utility of aptamer as bio-recognition element, providing the high binding af-
finity and specificity against the target analytes. Recent research has focused on the development of
structure switching signaling aptamer assay, transducing the aptamer- target recognition event into an
easily detectable signal. In this paper, we demonstrate the development of structure switching aptamer
assay for determination of aflatoxin M1 (AFM1) employing the quenching-dequenching mechanism.
Hybridization of fluorescein labelled anti-AFM1 aptamer (F-aptamer) with TAMRA labelled complementary
sequences (Q-aptamer) brings the fluorophore and the quencher into close proximity, which
results in maximum fluorescence quenching. On addition of AFM1, the target induced conformational
formation of antiparallel G-quadruplex aptamer-AFM1 complex results in fluorescence recovery. Under
optimized experimental conditions, the developed method showed the good linearity with limit of detection
(LOD) at 5.0 ng kg1 for AFM1. The specificity of the sensing platform was carefully investigated
against aflatoxin B1 (AFB1) and ochratoxin A (OTA). The developed assay platform showed the high
specificity towards AFM1. The practical application of the developed aptamer assay was verified for
detection of AFM1 in spiked milk samples. Good recoveries were obtained in the range from 94.40% to
95.28% (n¼3) from AFM1 spiked milk sample