Preparation of an anti-thiamethoxam monoclonal antibody for development of an indirect competitive enzyme-linked immunosorbent assay and a colloidal gold immunoassay

Figures & data

Figure 1. Hapten synthesis of thiamethoxam.

Figure 2. The ultraviolet-visible absorption spectra of artificial antigen: (a) TMX-EDC-KLH; (b) TMX-EDC-OVA.

Figure 3. The indirect competitive inhibition non-linear standard curve for thiamethoxam of 4D2.

Table 1. Cross-reactivity of mAb to thiamethoxam and analogs.

Chemicals	Structure	IC50 (ng/mL)	CR (%)
thiamethoxam		0.87	100
clothianidin		>1000	<0.087
Thiacloprid		>1000	<0.087
Imidacloprid		>1000	<0.087
nitenpyram		>1000	<0.087
acetamiprid		>1000	<0.087

Table 2. Accuracy evaluation of immunoassay methods for the sample.

Sample	Spiked level (ng/g)	ic-ELISA			Strip
		Mean ± SD (ng/mL)	Recovery (%)	CV (%)	Control-line	Test-line
Cucumbers	5	0.484 ± 0.056	97 ± 11.2	11.57	+++	++
	10	0.894 ± 0.071	89 ± 7.1	7.94	+++	++
	25	2.332 ± 0.084	93 ± 3.4	3.60	+++	+
	50	4.598 ± 0.125	92 ± 2.5	2.72	+++	+
	100	9.337 ± 0.407	93 ± 4.1	4.40	+++	–

Figure 4. Optimisation of different antibodies and coating antigens. Antibody 4D2 and coating 2 were better. 1 = 0 ng/mL, 2 = 10 ng/mL.

Figure 5. The sensitivity of the immunochromatographic assay in PBS for thiamethoxam. 1 = 0 ng/mL, 2 = 0.5 ng/mL, 3 = 1 ng/mL b, 4 = 2.5 ng/mL, 5 = 5 ng/mL, 6 = 10 ng/mL. Cut off at 10 ng/mL.

Figure 6. The sensitivity of the immunochromatographic assay of cucumber sample for thiamethoxam.1 = 0 ng/mL, 2 = 0.5 ng/mL, 3 = 1 ng/mL b, 4 = 2.5 ng/mL, 5 = 5 ng/mL, 6 = 10 ng/mL. Respectively cut off at 2 and 1 ng/mL.