Methods and materials for the detection of dengue virus infection
Inventors
Raychaudhuri, Syamal • Houghton, Raymond L. • Morkowski, Stanislaw • Stevens, Yvonne • Bose, Mohua • Needham, James William • CHEN, Hongjing
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-8920804-B2
Publication Date
2014-12-30
Expiration Date
Abstract
The present invention provides monoclonal antibodies that are specific for the Dengue non-structural glycoprotein NS1 in monomeric and/or oligomeric (primarily dimeric) form, together with methods, including ELISA and lateral flow assays, that employ the disclosed antibodies for the early detection of Dengue virus infection. Diagnostic kits for the detection of Dengue infection are also provided, such kits including the disclosed monoclonal and/or polyclonal antibodies.
Core Innovation
The invention relates to dengue virus infection detection using dengue NS1 glycoprotein forms, specifically NS1 monomeric form and NS1 oligomeric form, and to monoclonal antibody and polyclonal antibody components that are specific for dengue NS1 polypeptide. It provides monoclonal antibodies or antigen binding fragments defined by heavy and light chain variable regions, and diagnostic formats that use these antibodies to detect dengue virus infection through NS1 binding and antigen detection interactions.
The invention addresses detection beyond existing RT-PCR and IgM ELISA by using antibody-based recognition of dengue NS1 polypeptide, including detection workflows intended for early detection. It describes assay formats consistent with sandwich ELISA and lateral flow assays, including combinations of capture antibody and detection antibody and labeled detection reagents that provide a measurable signal when dengue NS1 polypeptide is present in a biological sample.
In addition, the invention provides diagnostic kits and dipsticks for dengue virus infection detection, including components such as controls, buffers, and lysing buffer, and a lateral flow membrane layout with defined test and control lines or zones. The disclosed assay system uses antibody interactions with NS1 polypeptide forms, reporter groups, and immobilized control polypeptide regions to support pan-serotype reactivity and comparative performance between antibodies such as mAb 528.292 and reference antibody 7E11.
Claims Coverage
The provided independent claim coverage includes one independent claim directed to a specific monoclonal antibody or antigen binding fragment. The dependent claims refine claim scope into inventive features covering additional binding specifications to NS1 forms and detection formats, including characterization of detection reagent reporter groups, selected biological sample matrices, and a lateral flow dipstick structure with an immobilized control polypeptide zone. Across the independent claim and its refinements, the coverage centers on antibody specificity to a dengue NS1 polypeptide and NS1 detection workflows using antibody complexes and labeled detection reagents.
Monoclonal antibody specific for dengue NS1 polypeptide
A monoclonal antibody, or antigen binding fragment thereof, that is specific for a dengue virus NS1 polypeptide, wherein the monoclonal antibody comprises a heavy chain variable region comprising SEQ ID NO: 2 and a light chain variable region comprising SEQ ID NO: 3.
Dengue infection detection using monoclonal-polyclonal conjugate detection
A method detects dengue virus infection in a biological sample by forming monoclonal antibody-NS1 polypeptide conjugates using a monoclonal antibody, forming a further complex with a dengue NS1-specific polyclonal antibody, and detecting the resulting conjugate.
Selection of reporter group for detection reagent
The method characterizes the detection reagent by selecting the reporter group from enzymes, substrates, cofactors, inhibitors, dyes, radionuclides, luminescent groups, fluorescent groups, colorimetric indicators, and biotin.
Biological sample matrix selection
The method further specifies the biological sample is chosen from blood, serum, plasma, saliva, cerebrospinal fluid, or urine.
Lateral flow dipstick with test and immobilized control zones
A dengue virus infection detecting dipstick uses a lateral flow membrane with antibody-labeled sample and detection zones and an immobilized control polypeptide zone.
Binding to non-hexameric NS1 prior to detection
A method detects dengue virus infection in a biological sample by contacting the sample with a monoclonal antibody that binds non-hexameric NS1 to form a monoclonal antibody-NS1 polypeptide complex, then contacting the complex with a detection reagent that binds the complex to detect the infection.
Overall, the claim coverage is anchored by a monoclonal antibody defined by SEQ ID NO: 2, the heavy chain variable region, and SEQ ID NO: 3, the light chain variable region, that is specific for dengue virus NS1 polypeptide. The dependent inventive features further specify an NS1 detection workflow using monoclonal-polyclonal conjugate detection, define options for the detection reagent reporter group, restrict biological sample types, define a dipstick lateral flow membrane with a test and immobilized control polypeptide zone, and constrain detection to binding to a non-hexameric form of NS1 before detection.
Stated Advantages
Early detection relative to existing RT-PCR and IgM ELISA is described in the background/rationale.
Pan dengue serotype reactivity is described.
Comparative performance is described, including increased sensitivity for mAb 528.292 versus reference antibody 7E11.
Documented Applications
Detection of dengue virus infection in a biological sample using antibody-based assay formats including sandwich ELISA and lateral flow assays (dipsticks).
Use of diagnostic kits and dipsticks with a lateral flow membrane layout including test and control lines or zones for dengue NS1 polypeptide detection.
Interested in licensing this patent?