Development of indirect competitive enzyme-linked immunoassay of colistin for milk and egg analysis

Figures & data

Figure 1. Structural formulas of polymyxin B, colistin, and colistin methanesulphonate.

Figure 2. Scheme of synthesis conjugated antigens in carbodiimide condensation method (A) and by reductive amination of periodate-oxidazed glycoprotein (B).

Figure 3. Spectra of BSA and BSA-PMB×50(edc) solutions (0.1 mg/mL) treated with TNBS (A). Calibration line for the determination of free amino groups (B).

Table 1. The specificity and sensitivity of ELISAs based on heterologous antigens.

Analyte	Gel(pi)-PMB×10		Gel(pi)-COL×50
	IC50 (ng/mL)	CR (%)	IC50 (ng/mL)	CR (%)
PMB	5.0	100	16.8	100
COL	5.7	87.7	9.2	182.6
CMS	>1000	>0.5	>1000	>0.5

Figure 4. Interaction of anti-BSA-PMB×50(edc) with Gel(pi)-PMB×10 in various matrices. Each column represents the average value (n = 4), the error value corresponds to the standard deviation.

Figure 5. Calibration curves for determining the COL in milk (1% SM) and eggs (PBS-t). Each point represents the average value (n = 3), the error value corresponds to the standard deviation.

Table 2. Recovery rate of COL from milk and egg samples fortified with antibiotic at threshold content.

Matrix	Dilution factor	Concentration COL, ng/mL		Recovery rate (%)
		Fortified	Determined
Milk	10	100	100 ± 8.2*	100 ± 8.2*
		50	47 ± 3.5	94.7 ± 7
		25	26 ±3.9	104 ± 16
Egg	50	600	580 ± 7.9	96.7 ± 13
		300	271.7 ± 27.5	90.6 ± 9.2
		150	133 ± 2.9	88.9 ± 1.9

*Average value (n = 4) ± standard deviation.The bold values are established MRL values.