Ridascreen ochratoxin a 30 15

For the quantification of AFB1, an immunoenzymatic test in a competitive format was performed according to the enclosed instructions of the commercial test kit (RIDASCREEN Aflatoxin B1 30/15; Art. No. TR1211, R-Biopharm, Darmstadt, Germany).
For OTA quantification, the competitive enzyme immunoassay kit RIDASCREEN Ochratoxin A 30/15, R1312 (R-Biopharm, Darmstadt, Germany) was used following the manufacturer’s instructions.
FOR ZEN determination, the RIDASCREEN Zearalenon (Art. Nr.: R1401) kit was applied, following the instructions of the manufacturer.
The standard curves in the enzyme immunoassays, AFB1, OTA, and ZEN were obtained with the mean values of each of the six duplicated concentration levels: 0, 1, 5, 10, 20, and 50 μg L⁻¹; 0, 0.03, 0.1, 0.3, 1, and 3 μg L⁻¹; and 0, 0.05, 0.15, 0.45, 1.35, and 4.05 μg L⁻¹ respectively. Mycotoxin quantification was achieved using the software RIDASOFT.Win.net. The calculation performed in double determinations used a cubic spline function. According to the manufacturer, the limit of detection of the enzyme immunoassays was 1 μg kg⁻¹ for AFB1, 0.8 μg kg⁻¹ for OTA, and 1.75 μg kg⁻¹ for ZEN.

For determination of ochratoxin A, RIDASCREEN^® Ochratoxin A 30/15 (R-Biopharm AG, Darmstadt, Germany) was conducted as a competitive ELISA. The method was carried out according to the manufacturer’s guidelines [24 ].

Acetonitrile and methanol (both HPLC grade) were purchased from Carlo Erba Reagents (Milan, Italy). Dichloromethane and hydrochloric acid (both HPLC grade) were purchased from Mallinckrodt Baker (Milan, Italy). Ultrapure water was produced by a Millipore Milli-Q system (Millipore, Bedford, MA). Ochratoxin A (OTA), sodium chloride (NaCl, ACS grade), sodium hydrogen carbonate (NaHCO₃, ACS grade), Tween 20, and acetic acid were purchased from Sigma-Aldrich (Milan, Italy). OTA immunoaffinity columns (OchraTest™) were obtained from VICAM, A Waters Business (Milford, MA). Glass microfiber filters (GF/A) and paper filters (No. 4) were purchased from Whatman (Maidstone, UK). OTA immunoaffinity columns (OchraPrep^®) and ELISA test kits (RIDASCREEN^® Ochratoxin A 30/15) were purchased from R-Biopharm (R-Biopharm AG, Darmstadt, Germany).