Mc lf

Mobile phases were prepared from LC-MS-grade acetonitrile (Fisher Optima, ThermoFisher, Greater London, UK) and water used for LC-MS was obtained in-house. Sample preparation reagents were HPLC grade. Toxin standards used for preparation of calibration solutions (MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR, MC-HilR, MC-HtyR, MC-LR, [Asp3] MC-LR and Nodularin) were all obtained from Enzo Life Sciences, Exeter, UK. A certified standard of [Dha7] MC-LR was obtained from the Institute of Biotoxin Metrology, National Research Council Canada (NRCC, Halifax, NS, Canada). Reference standards received as solid films were dissolved in 50% aqueous methanol, to form stock solutions. A mixed stock solution was subsequently prepared by combining aliquots of each stock, followed by further dilutions to create seven-level suite of working calibration standards between 0.33 ng/mL to 327 ng/mL per toxin.

MC-LR, MC-RR, and MC-LA were purchased from Cayman Chemical (Ann Arbor, MI, USA). MC-YR, MC-LF, and MC-LW were purchased from Enzo Life Sciences (Farmingdale, NY, USA). C₂D₅ MC-LR was purchased from Cambridge Isotope Laboratories (Tewksbury, MA, USA). MC-LR-Cys was synthesized by the Westrick group (Wayne State University, Detroit, MI, USA) [46 (link)]. ACS-grade zinc sulfate (ZnSO₄) and HPLC-grade water, methanol (CH₃OH), and acetonitrile (CH₃CN) were purchased from Fisher Scientific (Pittsburgh, PA, USA). Reagent-grade formic acid (FA) was purchased from Sigma (St. Louis, MO, USA).

To investigate the selectivity of MC10E7 for other microcystins, standard dilutions of MC‐LW, MC‐LF and MC‐RR (all microcystins were purchased from Enzo Life Sciences, Lörrach, Germany) were prepared in methanol/water (1/10, v/v) to final concentrations in the range 0.001–1000 μg/L. Cross‐reactivities were calculated using the quotient of the IC₅₀ values (concentration giving 50% inhibition) relative to MC‐LR in per cent.

MCs (MC-LR, MC-YR, MC-RR, MC-LA, MC-LY and MC-LF) and NOD standards were purchased from Enzo Life Sciences, Ltd. (UK), CYN was obtained from n’Tox (France) and ATX-A was purchased from the National Research Council, Canada. MMPB was purchased from Wako Pure Chemical Industries, Ltd. (Japan). CYN N¹⁵-labelled internal standard (N¹⁴ → N¹⁵) was a gift from the Federal Institute for Materials Research and Testing (BAM), Berlin, Germany. Acetonitrile and methanol, LC-MS Chromasolv grade, dichloromethane, formic acid, trifluoroacetic acid, potassium permanganate, sodium (meta) periodate and sodium bisulfite were all purchased from Sigma-Aldrich, UK, as were the ENVI-Carb solid-phase extraction (SPE) cartridges (250 mg, 3 cm³). Oasis HLB and PRiME SPE cartridges (60 mg, 3 cm³) were purchased from Waters, Ireland. The water used was supplied from an in-house Milli-Q water system (Millipore, Ltd., UK), with conductivity and total organic content (TOC) of the water being 18 MΩ and 3 ppb, respectively.

The solvents for the extraction and for the basis of the mobile phase were UHPLC-MS grade. All toxin standards for MCs (MC-LR, MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR) and NOD came from Enzo Life Sciences^® (Antwerp, Belgium) and were received under the form of a solid powder. After dilution in 100% methanol (MeOH), mixed stock solutions were prepared in 50% methanol (MeOH) (50% Milli-Q water with 1% acetic acid (v/v)). The stock and the intermediate solutions were stored at −20 °C.

MC-LR and MC-RR certified standards, the anti-adda antibody (AD4G2), and the assay kit for detection of MCs/NOD in water by PP2A were purchased from Abraxis, Inc (Warminster, PA, USA). MC-LF, NOD, cantharidin, okadaic acid, and calyculin A standard were purchased from Enzo (Farmingdale, NY, USA). Norcantharidin and phosphate buffered saline, pH 7.4 with Tween 20 (PBS-T) were obtained from Sigma-Aldrich (St. Louis, MO, USA). All MCs and other PP2A inhibitors were stored at −20 °C. The antibody was stored at 4 °C. Acetonitrile was from Pharmaco (Dawsonville, GA, USA). Methanol and formic acid were obtained from Fisher Scientific (Waltham, MA, USA). Life Technologies Corporation (Grand Island, NY, USA) supplied the Dynabeads MyOne Streptavidin T1, Zebaspin 7K molecular weight cut off, 0.5 mL desalting columns, and EZ-link NHS-PEG4-biotin. The Thermomixer C, Protein LoBind 2 mL microcentrifuge tubes, Protein LoBind 1 mL deep well plates, and TwinTec Lo-Bind PCR plates were purchased from Eppendorf (Hauppage, NY, USA). V&P Scientific (San Diego, CA, USA) supplied the 96-well plate magnet (part number VP771HH-MC). The microcentrifuge tube magnet was purchased from Invitrogen (Waltham, MA, USA).

LC-MS grade acetonitrile, water and formic acid and HPLC-grade methanol and water were purchased from Fisher (ThermoFisher, UK). Reference toxin standards of microcystins (MC-LR, MC-RR, MC-YR, MC-WR, MC-LW, MC-LA, MC-LY, MC-LF, MC-HtyR, MC-HiLR, [Asp3]-MC-LR/[Dha7]-MC-LR) and NOD ≥95% were as per Enzo Life Sciences (Exeter, UK) (Fig. S1). A certified standard of [Dha7]-MC-LR and a pre-certified freeze-dried matrix reference material of blue-green algae (RM-BGA, Lot 201301) containing a range of MCs were purchased from the Institute of Biotoxin Metrology, National Research Council Canada (Ontario, Canada).
A mixed stock solution was prepared by combining aliquots of each toxin to give a final concentration of 327 μg/L. For external calibration, a seven point calibration curve was prepared by serial dilution with methanol/water (1:1, v/v) in the range of 0.33–327 μg/L for each toxin and stored at – 18 °C. A quality control reference material (RM-BGA, National Research Council, Halifax, Canada) was prepared, with toxins extracted in the supernatant after 28 mL of methanol/water (1:1, v/v) + 0.1% acetic acid were added to RM-BGA (280 mg) and subsequent centrifugation (4500  $\times$ g; 10 min).
Shellfish diet 1800 (approximately 7.4 × 10¹¹ cells/mL) was purchased from ReedMariculture Inc., (US) and dilutions were made in water/seawater (10:0.86, v/v).