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Label-free detection technique based on impedance was investigated for aflatoxin M1 (AFM1) and aflatoxin M2 (AFM2) analysis in milk products. The impedance change resulting from antigen-antibody interaction was studied using a two-electrode setup made up of silver (Ag) wire. Processed milk such as drinking yogurt and flavored milk samples were analyzed in a flow-based setup. Two microflow pumps were used to construct the flow system where analytes (AFM1 and AFM2) were injected and impedance was measured using functionalized Ag wire electrodes. The flow system was optimized by adjusting both inlet and outlet flows to maintain the reaction volume optimum for impedance measurements. Using Bode plot, the matrix effect was investigated for detection of AFM1 and AFM2 in various matrices. Good recoveries were obtained even at low-AFM1 concentrations in the range of 1-100 pg/mL. The influence of AFM2 on the detection of AFM1 was also investigated. The proposed method provides good scope for online monitoring of such hazardous toxins in milk products.

An ultra-sensitive sandwich ELISA was developed for detection of AFM1 in milk. The assay involved the immobilization of rat monoclonal antibody of AFM1 in 384 microtiter plate to capture AFM1 antigen. This was detected by tracer secondary rabbit poly-clonal antibody labelled with horseradish peroxidase upon addition of a luminol-based substrate. Milk samples with different fat percentage were analyzed after pre-treatment. Linear range of AFM1 detection 250-6.25 pg/mL was achieved in 3% fat milk. The miniaturised assay (10 μL) enabled ultra trace analysis of AFM1 in milk with much improved lower limit of detection at 0.005 pg/mL. A sensitive magnetic nanoparticles (MNPs) based ELISA was also developed and coupled with micro plate ELISA for analysis in milk. The hybrid-assay, by coupling the 1°Ab immobilized MNPs column with microwell plate assay enabled simultaneous measurement of low (0.5 pg/mL) and high AFM1 contamination (200 pg/mL). The most promising feature of this MNPs-ELISA is the small column size, high capture efficiency and lower cost over other reported materials. The proposed assay can be deployed for simultaneous analysis and monitoring of AFM1 in milk., (Copyright © 2010 Elsevier B.V. All rights reserved.)

A survey of aflatoxin M1 (AFM1) contamination in packaged milk and infant formula milk samples in the Goan market, India, was conducted using high performance liquid chromatography, association of analytical communities approved commercial kit and a sensitive chemiluminescent sandwich enzyme-linked immunosorbent assay (ELISA). A total of 72 samples of infant formula milk food (18) and packaged milk samples (54) was analysed. One hundred per cent of the analysed samples exceeded the European Communities recommended limits (50 ng/L) and 75% of the samples exceeded Codex Alimentarius, Food Safety and Standards Authority of India (FSSAI) and US Food and Drug Administration recommended limits (500 ng/L). The range of contamination of AFM1 was found lower in infant milk formula (501–713 ng/L) than liquid milk (511–809 ng/L). The methods were also compared for their performance, and ELISA was found to be most suitable for analysis of low-level AFM1 contamination in milk. [ABSTRACT FROM AUTHOR]

A sensitive fluorimetric ELISA was developed for the analysis of aflatoxins. The assay was performed in a 384 microwell plate, wherein high specificity monoclonal antibody against AFM1 (mAb-AFM1) was used as capture antibody and FITC conjugated secondary antibody was used for detection and quantification of the analyte. The linear range of the immunoassay was found to be 6.25-50 pg/mL. AFM1 as low as 1 pg/mL was detected by this method with assay volume 40 µL. The multi-analysis of different aflatoxins was also investigated in the microwell plate, based on the cross-reactivity (CR) approach. Real milk samples were tested along with certified reference material by standard addition method and recovery analysis was done. The mAb-AFM1 showed 23.2% CR with AFB1, 50% CR with respect to AFM2, and least CR towards AFG1 (<1%). Furthermore, mixture analysis of AFM2 and AFB1 was carried out at specific concentrations of AFM1. The advantages of this developed immunoassay are high sensitivity, high throughput, multianalyte detection, versatility, and ease of handling. [ABSTRACT FROM AUTHOR]

Abstract: A highly sensitive and selective label-free impedimetric immunosensor based on silver (Ag) wire electrode for the detection aflatoxin M1 (AFM1) in milk is presented. The sensor was constructed by functionalizing Ag wire coupled with selective monoclonal antibodies of AFM1 through self assembled monolayers. The antigen–antibody interaction was quantified by measuring impedance in the frequency range (1–100Hz) at 10mV applied ac potential. A linear working range (6.25–100pgmL−1) for AFM1 was obtained with 20min analysis time. The sensor sensitivity was found to be about 2.1% impedance change per decade with limit of quantitation 6.25pgmL−1 and limit of detection 1pgmL−1. Good recoveries were obtained with SD=1.13 and R 2 =0.982 in spiked milk samples. The proposed method is useful for sensitive analysis of AFM1 in milk. [Copyright &y& Elsevier]