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US 12,754,189 B2Granted
Drug-specific pharmacokinetic assay for IL-15 superagonist
AssigneeNantCell, Inc.
InventorsShiho Tanaka, Kayvan Niazi
Life of this patent
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Abstract
Provided herein, are methods and compositions for detecting heteromeric protein complexes in biological samples. The methods and compositions allow for capturing and detecting the protein complex with substantially the same antibody, while avoiding detection of native proteins.
Claims14
A composition comprising:
- (i) an interleukin-15 (IL-15)/interleukin-15 receptor alpha Sushi domain (IL-15RαSu) complex comprising:
- a) a first IL-15RαSu domain comprising SEQ ID NO:1;
- b) a second IL-15RαSu domain comprising SEQ ID NO:1, wherein the first and second IL-15RαSu domains are directly or indirectly joined by a disulfide bond;
- c) a first IL-15 domain, bound by electrostatic interactions to the first IL-15RαSu domain to form a first IL-15/IL-15RαSu complex;
- d) a second IL-15 domain, bound by electrostatic interactions to the second IL-15RαSu domain to form a second IL-15/IL-15RαSu complex;
- (ii) a first anti-IL-15 monoclonal antibody (mAb) bound to an epitope on the first IL-15/IL-15RαSu complex, wherein the first anti-IL-15 monoclonal antibody is biotinylated for conjugation to a streptavidin-conjugated polymeric surface;
- and
- (iii) a second anti-IL-15 mAb bound to the second IL-15/IL-15RαSu complex, wherein the anti-IL-15 second mAb comprises a detection means selected from the group consisting of a fluorophore, a radioisotope, and an enzyme;
- wherein both the first anti-IL-15 mAb and the second anti-IL-15 mAb comprise the same epitope binding domain.
6 more elements
A method for detecting a heterotetrameric IL-15/IL-15RαSu complex in a biological sample, the method comprising:
- a) contacting the biological sample comprising the IL-15/IL-15RαSu complex with a first anti-IL-15 mAb, wherein the anti-IL-15 mAb is biotinylated for conjugation to a streptavidin-conjugated polymeric surface, wherein the IL-15/IL-15RαSu complex comprises two IL-15 domains and two IL-15RαSu domains each comprising SEQ ID NO:1, wherein each IL-15 domain is electrostatically bound to an IL-15RαSu domain, wherein the two IL-15RαSu domains are directly or indirectly bound to each other by a disulfide bond, and wherein the Fab portion of the anti-IL-15 mAb binds an epitope on the IL-15/IL-15RαSu complex with an affinity between 500 nM and 1 fM;
- b) contacting the biological sample with a second anti-IL-15 mAb under conditions such that the second anti-IL-15 antibody binds to the identical epitope on the second IL-15/IL-15RαSu complex, wherein the second anti-IL-15 mAb comprises a detection means selected from the group consisting of a fluorophore, a radioisotope, and an enzyme;
- c) washing unbound complexes from the polymeric surface;
- and
- d) detecting binding of the second anti-IL-15 mAb.
2 more elements
Dependencies are inferred from the claim text.