A rapid competitive ELISA assay of Okadaic acid level based on epoxy-functionalized magnetic beads

Figures & data

Scheme 1. The rapid competitive ELISA assay based on the epoxy-magnetic beads protocol.

Figure 1. The morphology of bare magnetic beads and glycosylated beads, EDX profiles and Fourier transform infrared (FTIR) characterization (A) The SEM and EDX profiles of bare magnetic beads (B) The SEM and EDX profiles of glycosylated beads. (C) FTIR spectra of the glucose and glycosylated MBs samples: a-glucose b-glycosylated MBs.

Figure 2. Cross-reactivity under the coexistence of other interferences. The IC₅₀ investigation for OA ranged from 0.39 to 25.0 µg L⁻¹, and DTX-1 ranged from 0.78 to 50.0 µg L⁻¹. The assay condition of insert table consisted of the interfering substance at a concentration of 25.0 µg L⁻¹.

Figure 3. Calibration curves of the presented MB protocol, routine ELISA protocol.

Table 1. The analytical performance comparison of relevant works.

Number	Type	LOD (µg L^−1)	IC50 (µg L^−1)	Linear range (µg L^−1)	ELISA strategy
1	Immunosensor	0.15	/	0.19–25	Immunosensor with streptavidin-MBs Indirect competitive [Dominguez et al., 2012]
2	Immunosensor	0.05	/	0.2–20	Fluorescence immunosensor with streptavidin-MBs. Indirect competitive [Pan et al., 2018]
3	ELISA kit	0.45	4.4	0.4–12.5	Routine ELISA kit, Indirect competitive [Lu et al., 2012]
4	ELISA kit	1.0	/	1.0–90	ELISA with protein G-MBs. Direct competitive [Hayat et al., 2012]
5	ELISA kit	31.3 µg OA eq/kg^a	/	31.3–1703 µg OA eq/kg	Routine ELISA. Direct competitive [Johnson et al., 2016]
6	ELISA kit	0.8	3.92	/	ELISA with Streptavidin-MBs. indirect competitive [Hayat et al., 2011]
7	ELISA kit	0.35^b	3.27	0.35–25.0^c	ELISA with epoxy-MBs direct competitive [this work]

^aOA equivalents- OA eq.

^bEqually to 17.5 µg kg⁻¹ in 2.0 g of sample.

^cEqually to 17.5–1250 µg kg⁻¹.

Table 2. The spiking tests and recovery results (n = 4).

Item/number	Spiking level (50.0 µg kg^−1)		Spiking level (100.0 µg kg^−1)		Spiking level (300.0 µg kg^−1)
	Result (µg kg^−1)	Recovery Rate (%)	Result (µg kg^−1)	Recovery Rate (%)	Result (µg kg^−1)	Recovery Rate (%)
Averages ± SD^a	48.28 ± 5.36	96.56 ± 10.72	97.93 ± 7.81	97.93 ± 7.81	307.5 ± 15.7	102.5 ± 5.23
CVs	11.09		7.98		5.12

^aQuartic analysis for every spiking level.

Table 3. Comparison of real sample detection within the three kinds of approaches (n = 4).

Samples	Detection results (µg kg^−1) Mean ± SD^a
	MBs approach	Commercial ELISA Kits	HPLC-MS/MS
Sinonovacula Constricta −01	17.56 ± 3.24	N.D^b	15.46 ± 1.35
−02	N.D	N.D	8.81 ± 1.17
−03	N.D	N.D	N.D
Pectinid −01	N.D	N.D	11.45 ± 1.02
−02	N.D	N.D	N.D
−03	N.D	N.D	N.D
Mytilus Coruscus −01	54.67 ± 5.67	57.23 ± 6.24	46.37 ± 4.35
−02	32.17 ± 2.45	N.D	25.17 ± 2.74
−03	N.D	N.D	N.D

^aAnalysis for every real sample (n = 4).

^bNo detection.

Dominguez, R. B., Hayat, A., Sassolas, A., Alonso, G. A., Munoz, R., & Marty, J. L. (2012). Automated flow-through amperometric immunosensor for highly sensitive and on-line detection of Okadaic acid in mussel sample. Talanta, 99, 232–237. doi: 10.1016/j.talanta.2012.05.045

 PubMed Web of Science ®Google Scholar

Pan, Y., Wei, X., Liang, T., Zhou, J., Wan, H., Hu, N., & Wang, P. (2018). A magnetic beads-based portable flow cytometry immunosensor for in-situ detection of marine biotoxin. Biomedical Microdevices, 20, 3. doi: 10.1007/s10544-018-0304-6

 Web of Science ®Google Scholar

Lu, S.-Y., Lin, C., Li, Y.-S., Zhou, Y., Meng, X.-M., Yu, S.-Y., … Liu, Z.-S. (2012). A screening lateral flow immunochromatographic assay for on-site detection of Okadaic acid in shellfish products. Analytical Biochemistry, 422(2), 59–65. doi: 10.1016/j.ab.2011.12.039

 PubMed Web of Science ®Google Scholar

Hayat, A., Barthelmebs, L., Sassolas, A., & Marty, J.-L. (2012). Development of a novel label-free amperometric immunosensor for the detection of Okadaic acid. Analytica Chimica Acta, 724, 92–97. doi: 10.1016/j.aca.2012.02.035

 PubMed Web of Science ®Google Scholar

Johnson, S., Harrison, K., & Turner, A. D. (2016). Application of rapid test kits for the determination of diarrhetic shellfish poisoning (DSP) toxins in bivalve molluscs from Great Britain. Toxicon, 111, 121–129. doi: 10.1016/j.toxicon.2016.01.052

 PubMed Web of Science ®Google Scholar

Hayat, A., Barthelmebs, L., & Marty, J.-L. (2011). Enzyme-linked immunosensor based on super paramagnetic nanobeads for easy and rapid detection of Okadaic acid. Analytica Chimica Acta, 690(2), 248–252. doi: 10.1016/j.aca.2011.02.031

 PubMed Web of Science ®Google Scholar