High affinity monoclonal antibodies for detection of shiga toxin 2
Inventors
He, Xiaohua • Stanker, Larry H. • Skinner, Craig B.
Assignees
US Department of Agriculture USDA
Publication Number
US-9513287-B1
Publication Date
2016-12-06
Expiration Date
2033-09-27
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Abstract
High affinity monoclonal antibodies against Stx2f and hybridomas that produce such antibodies are described. The antibodies may be used in a kit for detecting Stx2f and variants thereof in a sample as well as neutralization of Shiga toxin in vivo.
Core Innovation
The invention describes high affinity monoclonal antibodies (Mabs) specific to Shiga toxin 2 (Stx2) and its variants, including the Stx2f subtype, as well as hybridoma cell lines producing these antibodies. These monoclonal antibodies can be used in methods and kits for detecting Stx2f and other Stx2 variants in samples such as patient, environmental, and food samples, and for neutralizing Shiga toxin in vivo.
The problem addressed is the difficulty in detecting Shiga toxin 2 due to its low concentration in complex matrices such as food, and the lack of commercially available antibodies with sufficient affinity and specificity for all Stx2 variants, particularly the divergent Stx2f subtype. Existing assays either lack sensitivity, specificity, or require complex procedures, making prompt diagnosis and outbreak control challenging.
To overcome these limitations, the patent provides monoclonal antibodies generated against genetically engineered toxoids or recombinant subunits of Stx2 and Stx2f, demonstrating high affinity (dissociation constants in the sub-nanomolar range), specificity for different Stx2 subtypes, suitability for sensitive immunoassay formats like sandwich ELISA and lateral-flow devices, and capability for in vivo neutralization and clearance of Stx2. These antibodies facilitate rapid, sensitive, and specific detection of Stx2 in complex samples and offer therapeutic potential.
Claims Coverage
The patent discloses three independent inventive features related to monoclonal antibodies against Shiga toxin 2f and methods or kits utilizing these antibodies for detection and capture of the toxin.
Monoclonal antibody produced by specific hybridoma cell lines
A monoclonal antibody produced by a hybridoma cell line selected from the group consisting of deposit accession numbers PTA-121022, PTA-121021, and PTA-122640.
Method for detecting Shiga toxin 2 using specific monoclonal antibodies
A method for detecting Shiga toxin 2 and its variants by incubating a sample with the monoclonal antibody produced by one or more of the specified hybridoma cell lines and detecting the antibody-toxin complex to indicate the presence or absence of Shiga toxin 2 in the sample.
Kit for detection of Shiga toxin 2 comprising specific monoclonal antibody
A kit for detecting Shiga toxin 2 in a sample that comprises: (1) a container containing monoclonal antibody produced by one or more of the hybridoma cell lines identified by deposit accession numbers PTA-121022, PTA-121021, and PTA-122640, and (2) instructions for using the antibody to bind Shiga toxin 2 and detect the immunological complex formation correlating with presence or absence of the toxin in the sample.
The claims cover monoclonal antibodies produced by specific hybridomas targeting Shiga toxin 2f, methods employing these antibodies for sensitive detection of the toxin in diverse samples, and kits comprising these antibodies with instructions, establishing tools for diagnostic and research applications against Stx2 and its variants.
Stated Advantages
Provides high affinity monoclonal antibodies with sub-nanomolar dissociation constants, enabling sensitive detection.
Enables rapid, sensitive, and specific detection of Shiga toxin 2 and variants in complex matrices like food, milk, and environmental samples.
Facilitates in vivo neutralization and clearance of Shiga toxin 2, offering potential therapeutic applications.
Preserves native toxin structure by using genetically engineered toxoids as immunogens, enhancing antibody binding to biologically active toxin.
Documented Applications
Use of monoclonal antibodies for rapid diagnostic tests detecting Stx2 and variants in patient, environmental, and food samples.
Development of sandwich ELISA and lateral-flow immunoassay formats for sensitive detection of Stx2a and Stx2f.
In vivo treatment and neutralization of Stx2 toxin exposure or infection in mammals using the monoclonal antibodies.
Use of monoclonal antibodies as vaccine or therapeutic agents to provide protection against Shiga toxin exposure.
Colony immunoblot assays for identifying Stx2f-producing E. coli in food samples, including poultry.
Capture and isolation methods for Shiga toxin 2 from complex samples using the monoclonal antibodies.
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