Development and validation of an immunochromatography test strip for rapid detection of pyrimethanil residues

Figures & data

Figure 1. Schematic drawing of the ICTS for pyrimethanil detection.

Table 1. A review of immunoassay methods for the determination of pyrimethanil in recent years.

Method	Antibody type	IC50 (ng/mL)	LOD (ng/mL)	Cut-off (ng/mL)	Sample	References
Ic-ELISA	Polyclonal antibody	1.1	0.024	Not detected	Carrot juice	(Mercader et al., 2012)
Ic-ELISA	Monoclonal antibody	0.22–12.3	0.04–0.05	Not detected	Strawberry, tomato and cucumber	(Esteve-Turrillas et al., 2015)
Ic-ELISA	Monoclonal antibody	0.25	0.02–0.04	Not detected	Grape, strawberry, peach, tomato, apple, and cucumber	(Cao et al., 2019)
Ic-ELISA	Monoclonal antibody	4	Not detected	Not detected	Cucumber	(Chen et al., 2019)
ICST	Monoclonal antibody	Not detected	Not detected	50	Cucumber	(Chen et al., 2019)
ICST	Monoclonal antibody	4.8	0.9	40	Grape, strawberry, peach, tomato, apple, and cucumber	This work

Figure 2. The effect of immobilization concentration and incorporation rates of coating antigen. Concentration of coating antigen (pMm–BSA) on the T1, T2, T3 and T4 were 1, 0.5, 0.25 and 0.125 mg/mL, respectively. Incorporation rates of pMm–BSA were 8:1, 16:1 and 32:1. The standard concentration: negative (0 ng/mL); positive samples (40 ng/mL).

Figure 3. Series of pyrimethanil standard samples tested by developed ICTS and visual results examined after 10 min. The standard solutions of pyrimethanil at each final concentration of 0, 0.625, 1.25, 2.5, 5, 10, 20, 40, and 80 ng/mL in sample (0.1 M PBS, pH 7.4).

Figure 4. Standard curve of the developed ICTS for pyrimethanil quantitation in PBS (n = 5). Competitive inhibition rate was defined as B/B₀ × 100%, where B₀ and B represent the ROD_T/ROD_C values of the sample with/without pyrimethanil, respectively. Typical calibration curves of the ICTS by the portable strip reader with increasing pyrimethanil concentrations, from top to bottom: 0, 0.625, 1.25, 2.5, 5, 10, 20, 40, and 80 ng/mL, respectively. Good linearity of the calibration curve was achieved for pyrimethanil in the range of 1.25–20 ng/mL.

Table 2. Accuracy and reproductivity of the ICTS in pyrimethanil-fortified samples.

Sample	Fortified conc. (μg/kg)	Intra-assay		Inter-assay^a
		Mean recovery^b (%)	CV (%)	Mean recovery^b (%)	CV (%)
Grape	5	96.3	6.3	103.6	7.5
	10	85.2	11.0	88.4	11.7
	50	85.9	10.3	83.3	9.8
Strawberry	5	85.7	10.7	91.5	11.2
	10	89.3	9.1	98.2	9.5
	50	91.7	7.8	96.5	8.8
Peach	5	89.8	8.5	93.6	9.2
	10	90.6	10.7	94.9	11.5
	50	89.1	9.1	97.1	9.9
Tomato	5	88.7	10.4	92.5	11.6
	10	90.0	9.3	98.6	9.7
	50	92.3	8.9	97.8	9.2
Apple	5	89.5	10.4	99.6	10.6
	10	93.7	8.1	98.8	9.6
	50	98.7	7.9	105.7	8.6
Cucumber	5	87.4	11.1	90.5	10.5
	10	89.6	8.8	94.7	9.7
	50	86.9	10.3	89.4	11.3

^aAssay was completed every 1 d for 3 days continuously.

^bMean value of five replicates at each fortified concentration.

Figure 5. Bland-Altman plots to show comparisons between detection results obtained by (a) the ICTS and ic-ELISA, and (b) the ICTS and HPLC-MS/MS in fortified fruit and vegetable samples.

Mercader, J. V., Esteve-Turrillas, F. A., Agullo, C., Abad-Somovilla, A., & Abad-Fuentes, A. (2012). Antibody generation and immunoassay development in diverse formats for pyrimethanil specific and sensitive analysis. The Analyst, 137(23), 5672–5679. doi: 10.1039/c2an35801h

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Esteve-Turrillas, F. A., Abad-Somovilla, A., Quinones-Reyes, G., Agullo, C., Mercader, J. V., & Abad-Fuentes, A. (2015). Monoclonal antibody-based immunoassays for cyprodinil residue analysis in QuEChERS-based fruit extracts. Food Chemistry, 187, 530–536. doi: 10.1016/j.foodchem.2015.04.119

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Cao, Y., Shi, H. X., Le, T., Tang, R., & Xie, Y. (2019). Development a monoclonal antibody based enzyme linked immunosorbent assay for screening pyrimethanil in fruits and vegetables. Food and Agricultural Immunology, 30(1), 548–563. doi: 10.1080/09540105.2019.1608160

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Chen, Z., Wu, X., Xu, L., Liu, L., Kuang, H., & Cui, G. (2019). Development of immunocolloidal strip for rapid detection of pyrimethanil. Food and Agricultural Immunology, 30(1), 1239–1252. doi: 10.1080/09540105.2019.1677566

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