title: Efficient Conjugation of Aflatoxin M1 With Bovine Serum Albumin through Aflatoxin M1-(O-carboxymethyl) Oxime and Production of Anti-aflatoxin M1 Antibodies.
abstract: BACKGROUND
  Aflatoxins are the most extensively studied group of mycotoxins produced by molds, especially the Aspergillus group, which are highly toxic to animals and humans.
  
  
  OBJECTIVE
  Since immunoassay is a simple and rapid method for the analysis of many toxic substances in comparison to the chromatographic methods, it is necessary to produce specific and sensitive antibodies for detection of Aflatoxin M1 (AFM1). The current study was conducted to produce bioconjugate of Aflatoxin M1 (AFM1) with Bovine Serum Albumin (BSA) as well as to generate specific antibodies against AFM1 for immunoassay of the mycotoxin.
  
  
  METHODS
  First, AFM1 was converted to AFM1-(O-carboxymethyl) oxime derivative. Then, AFM1-oxime was coupled with BSA and the product was assessed by UV-VIS spectrophotometry. In order to generate polyclonal antibodies against AFM1, rabbits were immunized with BSA-AFM1 conjugate. Produced antibodies were purified using ion exchange chromatography and BSA-Sepharose 4B affinity chromatography. The titers and specificity of the produced antibodies were determined by Enzyme-Linked Immunosorbent Assay (ELISA).
  
  
  RESULTS
  The results indicated that coupling of AFM1 with O-(Carboxymethyl) hydroxylamine hemihydrochloride was suitable and 12 moles of AFM1-oxime were successfully coupled to each mole of BSA. In addition, the titers and specificity of the prepared antibody were considerable compared to standard anti-AFM1 antibodies. The relative cross-reactivity of each toxin (relative to AFM1) with purified anti-AFM1 antibodies, as determined by the amount of aflatoxin necessary to cause 50% inhibition of enzyme activity, was 70, 105, 240, and 2500 ng/mL for AFB1, AFB2, AFG1, and AFG2, respectively.
  
  
  CONCLUSIONS
  The prepared antibody can be used for the development of an ELISA kit to assay AFM1 in milk and other biological fluids.
authors.affiliation: Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah, IR Iran.
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Khademi
authors.affiliation: Department of Biology, Faculty of Sciences, Payame Noor University, Tehran, IR Iran.
authors.forename: Masoud
authors.initials: M
authors.lastname: Mohammadi
authors.affiliation: School of Pharmacy, Kermanshah University of Medical Sciences, Kermanshah, IR Iran.
authors.forename: Amir
authors.initials: A
authors.lastname: Kiani
authors.affiliation: Department of Biology, Faculty of Sciences, Payame Noor University, Tehran, IR Iran.
authors.forename: Reza
authors.initials: R
authors.lastname: Haji Hosseini Baghdadabadi
authors.affiliation: Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah, IR Iran.
authors.forename: Shahram
authors.initials: S
authors.lastname: Parvaneh
authors.affiliation: Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah, IR Iran.
authors.forename: Ali
authors.initials: A
authors.lastname: Mostafaie
keywords: Aflatoxin M1; Bovine Serum Albumin; ELISA; Toxin Conjugate