title: Development and Validation of a Fluorescent Multiplexed Immunoassay for Measurement of Transgenic Proteins in Cotton (Gossypium hirsutum).
abstract: In order to provide farmers with better and more customized alternatives to improve yields, combining multiple genetically modified (GM) traits into a single product (called stacked trait crops) is becoming prevalent. Trait protein expression levels are used to characterize new GM products and establish exposure limits, two important components of safety assessment. Developing a multiplexed immunoassay capable of measuring all trait proteins in the same sample allows for higher sample throughput and savings in both time and expense. Fluorescent (bead-based) multiplexed immunoassays (FMI) have gained wide acceptance in mammalian research and in clinical applications. In order to facilitate the measurement of stacked GM traits, we have developed and validated an FMI assay that can measure five different proteins (β-glucuronidase, neomycin phosphotransferase II, Cry1Ac, Cry2Ab2, and CP4 5-enolpyruvyl-shikimate-3-phosphate synthase) present in cotton leaf from a stacked trait product. Expression levels of the five proteins determined by FMI in cotton leaf tissues have been evaluated relative to expression levels determined by enzyme-linked immunosorbent assays (ELISAs) of the individual proteins and shown to be comparable. The FMI met characterization requirements similar to those used for ELISA. Therefore, it is reasonable to conclude that FMI results are equivalent to those determined by conventional individual ELISAs to measure GM protein expression levels in stacked trait products but with significantly higher throughput, reduced time, and more efficient use of resources.
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Grant R
authors.initials: GR
authors.lastname: Yeaman
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Sudakshina
authors.initials: S
authors.lastname: Paul
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Iryna
authors.initials: I
authors.lastname: Nahirna
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Yongcheng
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Andrew E
authors.initials: AE
authors.lastname: Deffenbaugh
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Zi Lucy
authors.initials: ZL
authors.lastname: Liu
authors.affiliation: Monsanto Company , 800 North Lindbergh Boulevard, St. Louis, Missouri 63167, United States.
authors.forename: Kevin C
authors.initials: KC
authors.lastname: Glenn
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005453:Fluorescence; D003368:Gossypium / Q000737:chemistry* / Q000235:genetics; D030821:Plants, Genetically Modified / Q000737:chemistry* / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis*
chemical_list: D011994:Recombinant Proteins
keywords: ELISA; Gossypium hirsutum; Luminex; cotton; fluorescent; multiplexed immunoassay