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Antibody characterization and immunoassays for palytoxin using an SPR biosensor

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Abstract

Palytoxin (PLTX), a polyether marine toxin originally isolated from the zoanthid Palythoa toxica, is one of the most toxic non-protein substances known. Fatal poisonings have been linked to ingestion of PLTX-contaminated seafood, and effects in humans have been associated with dermal and inhalational exposure to PLTX containing organisms and waters. Additionally, PLTX co-occurrence with other well-characterized seafood toxins (e.g., ciguatoxins, saxitoxins, tetrodotoxin) has hindered direct associations of PLTX to seafood-borne illnesses. There are currently no validated methods for the quantitative detection of PLTX(s). As such, a well-characterized, robust, specific analytical technique is needed for the detection of PLTX(s) in source organisms, surrounding waters, and clinical samples. Surface plasmon resonance (SPR) biosensors are ideally suited for antibody characterization and quantitative immunoassay detection. Herein, we describe a newly developed SPR assay for PLTX. An anti-mouse substrate was used to characterize the kinetic values for a previously developed monoclonal anti-PLTX. The characterized antibody was then incorporated into a sensitive, rapid, and selective PLTX assay. Buffer type, flow rate, analyte-binding time, and regeneration conditions were optimized for the antibody–PLTX system. Cross-reactivity to potentially co-occurring seafood toxins was also evaluated. We show that this optimized assay is capable of measuring low- to sub-ng/mL PLTX levels in buffer and two seafood matrices (grouper and clam). Preliminary results indicate that this SPR biosensor assay allows for (1) rapid characterization of antibodies and (2) rapid, sensitive PLTX concentration determination in seafood matrices. Method development information contained herein may be broadly applied to future PLTX detection and/or antibody characterization efforts.

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Fig. 1
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Abbreviations

EFSA:

European Food Safety Authority

HBS-EP+:

10 mM HEPES, 150 mM NaCl, 3 mM EDTA, and 0.05% (v/v) surfactant P20 buffer

k a :

Association rate constant

K D :

Dissociation constant

k d :

Dissociation rate constant

PBS:

10 mM phosphate-buffered saline pH 7.4

PLTX:

Palytoxin

R max :

Analyte-binding capacity

RU:

Resonance unit

SPR:

Surface plasmon resonance

t c :

Mass transfer constant (units RU × M−1s−2/3m−1/3)

U :

Uniqueness value

χ 2 :

Average-squared residual

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Authors and Affiliations

  1. Center for Food Safety and Applied Nutrition, US Food and Drug Administration, 5100 Paint Branch Parkway, College Park, MD, 20740, USA

    Betsy Jean Yakes, Stacey L. DeGrasse & Jonathan R. Deeds

  2. Integrated Toxicology Division, US Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, MD, 21702, USA

    Mark Poli

Authors
  1. Betsy Jean Yakes
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  2. Stacey L. DeGrasse
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  3. Mark Poli
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  4. Jonathan R. Deeds
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Correspondence to Betsy Jean Yakes.

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Yakes, B.J., DeGrasse, S.L., Poli, M. et al. Antibody characterization and immunoassays for palytoxin using an SPR biosensor. Anal Bioanal Chem 400, 2865–2869 (2011). https://doi.org/10.1007/s00216-011-5019-y

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