Diagnostic assays and methods of use for detection of filarial infection
Inventors
Nutman, Thomas B. • Fink, Doran • Kubofcik, Joseph • Burbelo, Peter D.
Assignees
US Department of Health and Human Services
Publication Number
US-9068993-B2
Publication Date
2015-06-30
Expiration Date
2031-10-31
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Abstract
The polynucleotide encoding the antigen Wb123 from the filarial nematode Wuchereria bancrofti, the major causative organism of lymphatic filariasis is provided, along with the polypeptide encoded by the polynucleotide. Methods for making the WM23 antigen, recombinant vectors encoding the Wb123 polynucleotide, and methods of detection of the Wb123 antigen through luciferase immunoprecipitation, ELISA and other detection systems are also provided.
Core Innovation
Wuchereria bancrofti is the major causative organism of lymphatic filariasis (LF), infecting millions globally and posing a diagnostic challenge due to a prepatent period during which standard diagnostics are ineffective. Existing antigen-based assays such as those using Bm-14 suffer from significant cross-reactivity with related filarial parasites and other parasitic infections, complicating accurate detection of Wuchereria bancrofti and Brugia malayi infections.
The invention identifies a specific antigen, Wb123, from Wuchereria bancrofti infective larvae (L3) that provides a rapid, high throughput, and specific diagnostic tool for detecting infections by Wuchereria bancrofti and Brugia malayi. This antigen can be used not only for individual diagnosis but also for early detection of recrudescent infections during control efforts and for mapping new areas of transmission.
The invention includes isolated polynucleotides encoding the Wb123 antigen (SEQ ID NO: 1), polypeptides encoded by this sequence, recombinant expression vectors, host cells expressing Wb123, and immunoassay methods such as luciferase immunoprecipitation system (LIPS) and ELISA to detect antibodies specific to Wb123. These diagnostic methods demonstrate specificity with minimal cross-reactivity to other filarial and non-filarial infections.
Claims Coverage
The patent includes three main independent claims detailing methods for detecting antibodies to Wb123 and exposure to Wuchereria bancrofti or Brugia malayi, as well as quantification of specific antibodies using solid supports.
Method for detecting antibodies to Wb123 antigen using a fusion protein with a reporter domain and immobilized second binding reagent
A method comprising contacting a sample with a fusion protein having a reporter domain and a binding domain containing Wb123 antigen encoded by SEQ ID NO: 1 to form an immunocomplex, contacting this with an immobilized second binding reagent capable of simultaneous binding to the antibodies if present to immobilize the fusion protein, detecting immobilization to determine presence or concentration of antibodies, and correlating detection with infection by Wuchereria bancrofti or Brugia malayi.
Method for detecting exposure to Wuchereria bancrofti or Brugia malayi via a luciferase immunoprecipitation assay
A method involving obtaining a sample from a subject, contacting the sample with a Renilla luciferase fusion protein comprising Wb123 antigen encoded by SEQ ID NO: 1 to form an immunocomplex, binding this complex to protein A/G, washing unbound components, incubating with a luciferase substrate (e.g., colenterazine), measuring luminescence, and correlating luminescence with exposure to the parasites.
Method for quantifying specific antibodies to Wb123 antigen using solid supports with immobilized antigen and antibody
A method involving immobilizing Wb123 antigen encoded by SEQ ID NO: 1 at a first location and an antibody which forms a complex with a sample antigen at a second location on a solid support, contacting the biological sample to both locations under conditions for immunocomplex formation, washing unbound material, separately detecting formation of each immunocomplex with labeled agents, and correlating detection amounts to quantify specific antibodies in the sample.
The independent claims cover inventive methods centered on detecting antibodies to Wb123 antigen utilizing fusion proteins with reporter domains and immobilized binding reagents, luciferase-based immunoassays for exposure detection, and quantitative antibody detection using solid supports, all specifically employing the Wb123 antigen encoded by SEQ ID NO: 1 as the key diagnostic target.
Stated Advantages
Provides a rapid, high throughput, and specific diagnostic tool for Wuchereria bancrofti and Brugia malayi infections.
Enables early detection of recrudescent infections in areas undergoing control efforts and for mapping new transmission zones.
Demonstrates minimal cross-reactivity with other filarial parasites and non-filarial infections, improving diagnostic specificity.
Offers sensitive and point-of-care capable methods to detect infection prior to microfilariae appearance in blood.
Documented Applications
Diagnosis of individual Wuchereria bancrofti and Brugia malayi infections.
Early detection of recrudescent infections in areas of control, especially during mass drug administration for LF elimination.
Mapping of new areas of transmission of Wuchereria bancrofti and/or Brugia malayi infection.
Use in immunoassays including luciferase immunoprecipitation system (LIPS), ELISA, and related diagnostic platforms for detecting antibodies to the Wb123 antigen.
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