Palytoxin detection and quantification using the fluorescence polarization technique

Detection of palytoxin-like compounds by a flow cytometry-based immunoassay supported by functional and analytical methods

“…Under these conditions the Max/Min ratio was 54, which was deemed sufficient, and the IC 50 of the calibration curve was 1.83 ± 0.21 nM (4.91 ± 0.57 ng mL -1 ), which provided the maximum sensitivity with a LOD (evaluated trough IC 20 , as in previously published immunoassays [36,37]) of 0.47 ± 0.15 nM (1.27 ± 0.39 ng mL -1 ) ( Figure 3). This microsphere-based immunoassay for PLTX-like molecules performs similarly, in terms of sensitivity, to previously published immuno-detection methods using the same anti-PLTXmAb [28][29][30], and it is 100 times more sensitive than the fluorescence polarization receptorbased method [27]. This single, semi-quantitative screening tool is based on the use of a commercially available technology, Luminex, that offers the possibility of combining different single-detection assays, as those developed for some aquatic toxins [38][39][40], in multi-detection methods [33,34], which would allow saving precious amounts of sample and experimental time.…”

“…neuroblastoma cells or erythrocytes) have been used for the development of cytotoxicity assays [23][24][25] based on the capability of PLTX to transform the Na + /K + -ATPase into a non-specific ion channel, triggering a disturbance of the cellular ion homeostasis [26]. Also a spectroscopic technique has been developed based on the measurement of changes in fluorescence polarization by PLTX binding to fluorescently labeled Na + /K + -ATPase [27]. The generation of monoclonal and polyclonal anti-PLTX antibodies allowed the development of immunoassays using different techniques such as enzyme linked immunosorbent assay (ELISA), surface plasmon resonance (SPR) or electrochemiluminescence (ECL)-based sensors [28][29][30][31].…”

Warm Seawater Microalgae: Growth and Toxic Profile of Ostreopsis Spp. from European Coasts

“…The method to quantify the PLTX is based on the FP methodology. In several studies, the FP technology was used to quantify toxins [29,32,33] and recently it was applied to develop a sensitive PLTX detection method [28]. In general, the FP units of a Na,K-ATPase-F conjugate decreased in the presence of PLTX and/or PLTX-like compounds with the same mechanism of action.…”

“…This fraction was evaporated to dryness and solved in 500 µL of MeOH-H 2 O (1:1). This protocol previously described was extended with a filtration step, through a 10000 NMWL filter [28,29]. The filtered extract was diluted 1:1000 to the final volume of 250 µL to check the …”

“…Therefore, in this context, the aim of this work is to estimate the production of PLTX-like compounds in different cultures of Ostreopsis spp. growing in several biological conditions by using a Fluorescence Polarization (FP) technique recently developed [28].…”