Methods and compositions relating to anti-biotin antibodies
Inventors
Assignees
Publication Number
US-12103981-B2
Publication Date
2024-10-01
Expiration Date
2039-08-02
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Abstract
Increasingly, human subjects are taking biotin as a medical therapy or nutritional supplement. This practice causes difficulty in analysis of biological samples, such as blood samples, using biotin conjugate reagents since free biotin results in interference. Inventive antibodies, and antigen binding fragments thereof, which specifically bind to biotin of a biotin conjugate and which have a higher affinity for biotin of a biotin conjugate than for free biotin, are provided along with methods of their use.
Core Innovation
The invention provides isolated antibodies and antigen binding fragments which specifically bind to the biotin of a biotin conjugate, and which have a higher affinity for biotin when it is part of a conjugate than for free biotin. These antibodies may be monoclonal and can be labeled with a detectable marker or immobilized on a solid or semi-solid support. Specific sequences for heavy and light chain variable regions are disclosed, including variants and nucleotide-encoded forms.
The problem addressed is the difficulty in analyzing biological samples, such as blood, using biotin conjugate reagents due to interference from free biotin, particularly in individuals who consume biotin therapeutically or as a supplement. The inventive antibodies overcome this problem by showing little or no affinity for free biotin while retaining strong binding to biotin in conjugates.
Compositions include antibodies characterized by specific heavy and light chain variable region sequences or their variants, which may be purified or attached to solid supports. Methods involve using these antibodies in assays to detect biotin conjugates in biological samples, with minimized interference from free biotin. Immunoassay kits comprising these antibodies or hybridoma cells producing them are also provided.
Claims Coverage
There are three main inventive features claimed: the antibody with defined specificity and affinity, methods for detecting biotin conjugates using this antibody, and immunoassay kits incorporating the antibody or corresponding hybridomas.
Antibody specific for biotin of a biotin conjugate with higher affinity than for free biotin
An isolated monoclonal antibody or antigen binding fragment thereof that specifically binds to biotin of a biotin conjugate and has a higher affinity for biotin of a biotin conjugate than for free biotin. - The antibody comprises one of the following sets of variable regions: - Heavy chain variable region SEQ ID NO:1 with light chain variable region SEQ ID NO:2 - Heavy chain variable region SEQ ID NO:3 with light chain variable region SEQ ID NO:4 - Heavy chain variable region SEQ ID NO:5 with light chain variable region SEQ ID NO:6 - The antibody may further comprise an attached detectable label or be immobilized on a solid or semi-solid support. - The corresponding heavy and light chain regions may be encoded by SEQ ID NO:7 and SEQ ID NO:8; SEQ ID NO:9 and SEQ ID NO:10; or SEQ ID NO:11 and SEQ ID NO:12.
Hybridoma cell producing the claimed antibody
A hybridoma cell comprising a nucleic acid encoding the monoclonal antibody or antigen binding fragment as defined in the inventive feature above.
Method for detecting a biotin conjugate using the antibody
A method for detecting a biotin conjugate in a sample, comprising: 1. Contacting a sample containing or suspected of containing a biotin conjugate with the monoclonal anti-biotin antibody or antigen binding fragment defined above, under binding conditions. 2. Detecting binding of the antibody or fragment to the biotin conjugate. - The method may include ELISA, immunochromatography, antigen capture, flow cytometry, immunoblot, immunofluorescence, immunoprecipitation, immunodiffusion, competitive immunoassay, immunocytochemistry, radioimmunoassay, or combinations thereof. - Applicable to blood samples from human subjects who take biotin as a medical therapy or supplement.
Immunoassay kit comprising the antibody or hybridoma cell
An immunoassay kit comprising the monoclonal antibody or antibody binding fragment, or the hybridoma cell that produces it, as defined above.
The inventive features center on monoclonal antibodies with distinct specificity and affinity profiles for biotin conjugates over free biotin, the methods that use these antibodies for detection in samples, the production by hybridomas, and their implementation in immunoassay kits.
Stated Advantages
The inventive antibodies enable detection of biotin in biotin conjugates with no or minimal interference by free biotin, overcoming problems in analysis of biological samples from subjects who have consumed biotin.
These antibodies provide a solution for interference of free biotin in blood samples, an increasing problem for avidin or streptavidin-based assay methods used in medical diagnosis and related applications.
Documented Applications
Detection of biotin conjugates in biological samples, including blood samples from humans taking biotin as medical therapy or nutritional supplement, using immunoassays such as ELISA, immunochromatography, antigen capture, flow cytometry, immunoblot, immunoprecipitation, immunodiffusion, competitive immunoassay, immunocytochemistry, and radioimmunoassay.
Use in immunoassay kits for analysis of biological samples avoiding interference of free biotin, particularly for clinical or diagnostic applications.
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