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Fluorimetric immunoassay for multianalysis of aflatoxins.

Kanungo L, Bhand S - J Anal Methods Chem (2013)

Bottom Line: The linear range of the immunoassay was found to be 6.25-50 pg/mL.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.

View Article: PubMed Central - PubMed

Affiliation: Biosensor Lab., Department of Chemistry, BITS, Pilani-K. K. Birla Goa Campus, Goa 403726, India.

ABSTRACT
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  μ L. 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.

No MeSH data available.


Scheme of fluorimetric ELISA designed for analysis of AFM1.
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Related In: Results  -  Collection


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fig2: Scheme of fluorimetric ELISA designed for analysis of AFM1.

Mentions: In this work, a simple ELISA was developed using fluorimetric technique. We performed fluorimetric analysis of AFM1 using FITC conjugated secondary (2°) antibody in 384 microwell plate as shown in Figure 2. Multianalysis of different aflatoxins was carried out by CR approach. The microwell plate was coated with anti-AFM1 primary (1°) monoclonal antibody (mAb) which reacted with AFM1 and partly recognized structurally similar aflatoxins such as AFM2, AFB1, and AFG1. Subsequently we analyzed 2 real milk samples using standard addition method, and recovery rate was studied. The CR was studied and mixture analysis was carried out. The occurrence of cocontaminants was detected by this mixture analysis.


Fluorimetric immunoassay for multianalysis of aflatoxins.

Kanungo L, Bhand S - J Anal Methods Chem (2013)

Scheme of fluorimetric ELISA designed for analysis of AFM1.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3755385&req=5

fig2: Scheme of fluorimetric ELISA designed for analysis of AFM1.
Mentions: In this work, a simple ELISA was developed using fluorimetric technique. We performed fluorimetric analysis of AFM1 using FITC conjugated secondary (2°) antibody in 384 microwell plate as shown in Figure 2. Multianalysis of different aflatoxins was carried out by CR approach. The microwell plate was coated with anti-AFM1 primary (1°) monoclonal antibody (mAb) which reacted with AFM1 and partly recognized structurally similar aflatoxins such as AFM2, AFB1, and AFG1. Subsequently we analyzed 2 real milk samples using standard addition method, and recovery rate was studied. The CR was studied and mixture analysis was carried out. The occurrence of cocontaminants was detected by this mixture analysis.

Bottom Line: The linear range of the immunoassay was found to be 6.25-50 pg/mL.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.

View Article: PubMed Central - PubMed

Affiliation: Biosensor Lab., Department of Chemistry, BITS, Pilani-K. K. Birla Goa Campus, Goa 403726, India.

ABSTRACT
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  μ L. 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.

No MeSH data available.