title: Sensitive detection of active Shiga toxin using low cost CCD based optical detector.
abstract: To reduce the sources and incidence of food-borne illness there is a need to develop affordable, sensitive devices for detection of active toxins, such as Shiga toxin type 2 (Stx2). Currently the widely used methods for measuring Shiga toxin are immunoassay that cannot distinguish between the active form of the toxin, which poses a threat to life, to the inactive form which can bind to antibodies but show no toxicity. In this work, we determine toxin activity based on Shiga toxin inhibition of green fluorescent protein (GFP) combined with low cost charge-coupled device (CCD) fluorescence detection, which is more clinically relevant than immunoassay. For assay detection, a simple low cost fluorescence detection system was constructed using a CCD camera and light emitting diode (LED) excitation source, to measure GFP expression. The system was evaluated and compared to a commercial fluorometer using photomultiplier detection for detecting active Stx2 in the range 100 ng/mL-0.01 pg/mL. The result shows that there is a negative linear relationship between Stx2 concentrations and luminous intensity of GFP, imaged by the CCD camera (R(2)=0.85) or fluorometer (R(2)=0.86). The low cost (∼$300) CCD camera is capable of detecting Shiga toxin activity at comparable levels as a more expensive (∼$30,000) fluorometer. These results demonstrate the utility and the potential of low cost detectors for toxin activity; this approach may increase the availability of foodborne bacterial toxin diagnostics in regions where there are limited resources and could be readily adapted to the detection of other food-borne toxins.
authors: Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Albany, CA, United States. Electronic address: reuven.rasooly@ars.usda.gov.
authors: Reuven
authors: R
authors: Rasooly
authors: Division of Biology, Office of Science and Engineering, FDA, Silver Spring, MD 20993, United States; University of Maryland, College Park, MD 20742, United States.
authors: Josh
authors: J
authors: Balsam
authors: Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Albany, CA, United States.
authors: Bradley J
authors: BJ
authors: Hernlem
authors: Division of Biology, Office of Science and Engineering, FDA, Silver Spring, MD 20993, United States; Office of Cancer Complementary and Alternative Medicine, National Cancer Institute, Rockville, MD 20850, United States.
authors: Avraham
authors: A
authors: Rasooly
mesh_terms: D015374:Biosensing Techniques; D005504:Food Analysis; D049452:Green Fluorescent Proteins; D006801:Humans; D022641:Shiga Toxin 2 / Q000302:isolation & purification*
chemical_list: D022641:Shiga Toxin 2; D049452:Green Fluorescent Proteins
keywords: Adenovirus; CCD camera; Food; Shiga toxin