Screening of Paralytic Shellfish Posioning Toxins in Naturally Occurring Samples with Three Different Direct Competitive Enzyme-Linked Immunosorbent Assays

Chu, Fun Sun; Hsu, Kuo-Hui; Huang, Xuan; Barrett, Richard; Allison, Christine G.

doi:10.1021/jf960244w

Cited by 37 publications

(20 citation statements)

References 26 publications

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“…These results are fairly consistent with other immunoassay publications (Usleber et al, 2001 andDubois et al, 2010) however by use of a "cocktail" of antibodies, as described by Campbell et al (2011b) or two immunoassays (Chu et al, 1996), these difficulties may be overcome. Alternatively, the multiplexed nature of the microarray format could allow multiple separate tests to be run, enabling a comprehensive toxin profile assay.…”

Section: Resultssupporting

confidence: 91%

“…Often the sensitivity is high, but inaccurate results may occur as a result of the antibody specificity (Usleber et al, 2001;Dubois et al, 2010). One approach to alleviate the problem of underestimation of results is to combine two antibodies into the test format as described by Campbell et al (2011b) or employ two assays (Chu et al, 1996). Several commercial ELISA kits exist for the screening of PSTs, however only one has been applied to the determination of STX in water samples, the Abraxis Saxitoxin (PSP) ELISA kit.…”

Section: Introductionmentioning

confidence: 99%

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Development and validation of an ultrasensitive fluorescence planar waveguide biosensor for the detection of paralytic shellfish toxins in marine algae

Meneely

Campbell

Greef

et al. 2013

Biosensors and Bioelectronics

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Publisher rights This is the author's version of a work that was accepted for publication in Biosensors and Bioelectronics. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published in Biosensors and Bioelectronics, VOL 41, 03/2013 General rights Copyright for the publications made accessible via the Queen's University Belfast Research Portal is retained by the author(s) and / or other copyright owners and it is a condition of accessing these publications that users recognise and abide by the legal requirements associated with these rights. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting galley proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. ABSTRACTMarine dinoflagellates of the genera Alexandrium are well known producers of the potent neurotoxic paralytic shellfish toxins that can enter the food web and ultimately present a serious risk to public health in addition to causing huge economic losses. Direct coastal monitoring of Alexandrium spp. can provide early warning of potential shellfish contamination and risks to consumers and so a rapid, sensitive, portable and easyͲtoͲuse assay has been developed for this purpose using an innovative planar waveguide device.The disposable planar waveguide is comprised of a transparent substrate onto which an array of toxinͲ protein conjugates is deposited, assembled in a cartridge allowing the introduction of sample, and detection reagents. The competitive assay format uses a high affinity antibody to paralytic shellfish toxins with a detection signal generated via a fluorescently labelled secondary antibody. The waveguide cartridge is analysed by a simple reader device and results are displayed on a laptop computer. Assay speed has been optimised to enable measurement within 15 min. A rapid, portable sample preparation technique was developed for Alexandrium spp. in seawater to ensure analysis was completed within a short period of time. The assay was validated and the LOD and CCɴ were determined as 12pg/mL and 20pg/mL respectively with an intraͲassay CV of 11.3% at the CCɴ and an average recovery of 106%. The highly innovative assay was proven to accurately detect toxin presence in algae sampled from the US and European waters at an unprecedented cell density of 10cells/L.

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Section: Resultssupporting

confidence: 91%

Section: Introductionmentioning

confidence: 99%

Development and validation of an ultrasensitive fluorescence planar waveguide biosensor for the detection of paralytic shellfish toxins in marine algae

Meneely

Campbell

Greef

et al. 2013

Biosensors and Bioelectronics

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“…In these cases ELISA results compared well with other methods, although this was because the samples fortuitously possessed a congener profile that reacted with the antibody in a fashion that matched the potency. Good agreement between ELISAs and MBA was also reported in other studies (Chu et al, 1996;Usleber et al, 1997), but not in all (Kasuga et al, 1996). As stated earlier, toxin profiles can be quite complex, making the cross-reactivity challenge with antibodies an impediment to widespread use of the kits.…”

Section: Toxin Detection Methodssupporting

confidence: 64%

Paralytic shellfish poisoning: Seafood safety and human health perspectives

Etheridge

2010

Toxicon

275

145

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a b s t r a c tParalytic shellfish poisoning (PSP) is the foodborne illness associated with the consumption of seafood products contaminated with the neurotoxins known collectively as saxitoxins (STXs). This family of neurotoxins binds to voltage-gated sodium channels, thereby attenuating action potentials by preventing the passage of sodium ions across the membrane. Symptoms include tingling, numbness, headaches, weakness and difficulty breathing. Medical treatment is to provide respiratory support, without which the prognosis can be fatal. To protect human health, seafood harvesting bans are in effect when toxins exceed a safe action level (typically 80 mg STX eq 100 g À1 tissue). Though worldwide fatalities have occurred, successful management and monitoring programs have minimized PSP cases and associated deaths. Much is known about the toxin sources, primarily certain dinoflagellate species, and there is extensive information on toxin transfer to traditional vectors -filter-feeding molluscan bivalves. Non-traditional vectors, such as puffer fish and lobster, may also pose a risk. Rapid and reliable detection methods are critical for toxin monitoring in a wide range of matrices, and these methods must be appropriately validated for regulatory purposes. This paper highlights PSP seafood safety concerns, documented human cases, applied detection methods as well as monitoring and management strategies for preventing PSP-contaminated seafood products from entering the food supply.Published by Elsevier Ltd. Paralytic shellfish poisoning toxins and sourcesParalytic shellfish poisoning (PSP) is a common seafood toxicity problem with worldwide distribution, and typically this illness is due to the consumption of contaminated molluscan bivalves and other shellfish. A similar seafood-related syndrome involves puffer fish contaminated with the same family of toxins. To distinguish these puffer fish poisonings from those caused by tetrodotoxin, this food poisoning syndrome is becoming known in the literature as saxitoxin puffer fish poisoning (SPFP; Landsberg et al., 2006;Deeds et al., 2008a). The toxins responsible for both of these seafood-borne illnesses are the neurotoxins known collectively as the saxitoxins (STXs), also referred to as PSP toxins (or PSTs). At least 24 saxitoxin-like congeners have been identified (Fig. 1), with a range of hydroxyl, carbamyl, and sulfate moieties at four sites on the backbone structure. These substitutions result in congeners varying more than three orders of magnitude in potency (Oshima et al., 1993). The carbamate toxins are the most potent, and they include saxitoxin (STX), neosaxitoxin (NEO), and the gonyautoxins (GTX1-4). The decarbamoyl toxins (dcSTX, dcNEO, dcGTX1-4) have intermediate toxicity and are reported in certain bivalves, but are not commonly found in toxic dinoflagellates. The N-sulfocarbamoyl toxins (B1 [GTX5], B2 [GTX6] and C1-4) are less potent. There is a fourth group known as the deoxydecarbamoyl toxins, but their potency has not yet *

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“…PAb against saxitoxin (10 µg mL -1 , 50 µL) prepared in PGG-NaCl buffer [10] (0.01 mol L -1 piperazine-glycylglycine pH 10.0+NaCl 150 mmol L -1 ) were added to a microplate and left overnight at room temperature. After immobilization, the wells were washed three times with a solution of PBS-T (PBS+0.05% Tween 20).…”

Section: Direct Competitive Assaymentioning

confidence: 99%

“…Because of the highly specific antigen-antibody interaction, several laboratories have attempted to develop an immunoassay for PSP [9,10]. Although an ELISA kit [11] for saxitoxin is now on the market, free antibodies against this toxin are not commercially available, despite this toxin having been included in the list of chemical weapon compounds (Paris Convention 13/01/1993, Low 496 25 /11/1995 1A07) In this paper the production of specific antibodies against saxitoxin is described.…”

Section: Introductionmentioning

confidence: 99%

Production of antibodies and development of highly sensitive formats of enzyme immunoassay for saxitoxin analysis

et al. 2002

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In this paper the production of antibodies against saxitoxin (STX) is described, as is the optimization and comparison of two competitive ELISA formats (direct and indirect) for the detection of this toxin. Tests were performed in a 96-well microplate using the toxin-specific polyclonal antibodies produced in our laboratory, obtained from rabbits immunized with saxitoxin-keyhole limpet hemocyanin (STX-KLH). In indirect ELISA format saxitoxin, conjugated to bovine serum albumin (STX-BSA) was coated onto the microtitre plate and incubated with standard toxin and anti-STX antibody. A goat antirabbit IgG Peroxidase conjugate was used to enable detection. In the direct ELISA format, STX standard, STX conjugate to horseradish peroxidase (STX-HRP), and enzyme substrate/chromogen solution were sequentially added to the microplate after antibody coating.Results showed the saxitoxin detection limit to be 3 and 10 pg mL -1 for direct and indirect ELISA formats, respectively.The suitability of the assay for quantification of saxitoxin in mussels was also studied. Samples were spiked with saxitoxin before and after sample treatment to study the extraction efficiency and matrix effect, respectively. After treatment, samples were analysed at 1:1000 v/v dilution in PBS to minimize the matrix effect and to detect the regulatory limit of 40-80 µg saxitoxin per 100 g mussels as stipulated by the Food and Drug Administration. The efficiency of extraction of saxitoxin was from 72 to 102%. These data were confirmed by liquid chromatography coupled with fluorimetric detection, the technique currently used for quantitative determination of toxins in seafood.

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Screening of Paralytic Shellfish Posioning Toxins in Naturally Occurring Samples with Three Different Direct Competitive Enzyme-Linked Immunosorbent Assays

Cited by 37 publications

References 26 publications

Development and validation of an ultrasensitive fluorescence planar waveguide biosensor for the detection of paralytic shellfish toxins in marine algae

Development and validation of an ultrasensitive fluorescence planar waveguide biosensor for the detection of paralytic shellfish toxins in marine algae

Paralytic shellfish poisoning: Seafood safety and human health perspectives

Production of antibodies and development of highly sensitive formats of enzyme immunoassay for saxitoxin analysis

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