Center for Food Safety and Applied Nutrition, US Food and Drug Administration, College Park, MD 20740, USA.
Anal Bioanal Chem. 2011 Jul;400(9):2865-9. doi: 10.1007/s00216-011-5019-y. Epub 2011 Apr 27.
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.
海兔毒素（PLTX）是一种多醚海洋毒素，最初从共生藻 Palythoa toxica 中分离得到，是已知的最毒的非蛋白物质之一。摄入含有 PLTX 的海鲜会导致致命中毒，而人类则因接触含有 PLTX 的生物和水而出现皮肤和吸入暴露的影响。此外，PLTX 与其他特征明确的海鲜毒素（例如，雪卡毒素、石房蛤毒素、河豚毒素）同时存在，阻碍了将 PLTX 与食源性疾病直接联系起来。目前还没有定量检测 PLTX(s) 的方法。因此，需要一种经过充分验证的、强大的、特异性的分析技术，用于在源生物、周围水域和临床样本中检测 PLTX(s)。表面等离子体共振（SPR）生物传感器非常适合用于抗体表征和定量免疫分析检测。在此，我们描述了一种新开发的 PLTX SPR 分析方法。使用抗鼠基质来表征先前开发的抗 PLTX 单克隆抗体的动力学值。然后将表征的抗体纳入到敏感、快速和选择性的 PLTX 分析中。优化了缓冲液类型、流速、分析物结合时间和再生条件，以适应抗体-PLTX 系统。还评估了与潜在共存的海鲜毒素的交叉反应性。我们表明，这种优化的测定法能够测量缓冲液和两种海鲜基质（石斑鱼和蛤）中的低至亚纳克/毫升 PLTX 水平。初步结果表明，这种 SPR 生物传感器测定法允许（1）快速表征抗体和（2）快速、灵敏地确定海鲜基质中的 PLTX 浓度。本文中包含的方法开发信息可能广泛应用于未来的 PLTX 检测和/或抗体表征工作。
