Monoclonal antibodies specific for buprenorphine and metabolites thereof
View Patent ↗The present invention relates to monoclonal antibodies that specifically bind buprenorphine and/or at least one metabolic product thereof. The present invention further relates to buprenorphine metabolite conjugates for the production of monoclonal antibodies that specifically bind buprenorphine and/or at least one metabolic product thereof and hybridoma cells that produce the monoclonal antibodies. The invention also relates to immunoassay methods for determining buprenorphine and/or one or more buprenorphine metabolites in a sample using the novel antibodies and conjugates of the present invention.
1. A monoclonal antibody that specifically binds buprenorphine and that has less than 10% cross-reactivity to buprenorphine glucuronide, norbuprenorphine, and norbuprenorphine glucuronide.
2. A cell line that produced a monoclonal antibody that specifically binds buprenorphine and that has less than 10% cross-reactivity to buprenorphine glucuronide, norbuprenorphine, and norbuprenorphine glucuronide.
3. Hybridoma cell line BUP 2.2 having ATCC accession number PTA 7198 and producing an antibody according to claim 1 .
4. A method for determining buprenorphine in a sample comprising
a. forming a reaction mixture comprising the sample, a monoclonal antibody that binds specifically with buprenorphine with less than 10% cross-reactivity to buprenorphine glucuronide, norbuprenorphine, and norbuprenorphine glucuronide to form a buprenorphine-antibody complex, and an buprenorphine analog conjugate that competes with the buprenorphine in the sample to form a buprenorphine analog-antibody complex, wherein either the antibody or the analog conjugate is bound to a label that combines with either the buprenorphine or the antibody, respectively, to produce a detectable signal, and
b. determining the signal produced in step (a) as a measure of the buprenorphine in the sample.
5. The method of claim 4 wherein if the antibody is bound to the label, then the analog conjugate is bound to a solid phase, or if the analog conjugate is bound to the label, then the antibody is bound to a solid phase.
6. A monoclonal antibody that specifically binds buprenorphine glucuronide and that has essentially no cross-reactivity to buprenorphine, norbuprenorphine, and norbuprenorphine glucuronide.
7. A cell line that produces a monoclonal antibody that specifically binds buprenorphine glucuronide and that has essentially no cross-reactivity to buprenorphine, norbuprenorphine, and norbuprenorphine glucuronide.
8. Hybridoma cell like BUP 88.1.1 having ATCC accession number PTA 5023 and producing an antibody according to claim 6 .
9. A method for determining buprenorphine glucuronide in a sample comprising
a. forming a reaction mixture comprising the sample, a monoclonal antibody that binds specifically with buprenorphine glucuronide and that has essentially no cross-reactivity to buprenorphine, norbuprenorphine, and norbuprenorphine glucuronide to form a buprenorphine glucuronide-antibody complex, and a buprenorphine glucuronide analog conjugate that competes with the buprenorphine glucuronide in the sample to form a buprenorphine analog-antibody complex, wherein either the antibody or the analog conjugate is bound to a label that combines with either the buprenorphine glucuronide or the antibody, respectively, to produce a detectable signal, and
b. determining the signal produced in step (a) as a measure of buprenorphine glucuronide in the sample.
10. The method of claim 9 wherein if the antibody is bound to the label, then the analog conjugate is bound to a solid phase, or if the analog conjugate is bound to the label, then the antibody is bound to a solid phase.
11. A norbuprenorphine conjugate having the structure:
wherein X is a linking group consisting of 0–10 carbon atoms or heteroatoms including 0–2 substituted or unsubstituted aromatic rings, Y is O, NH, or S, and Z is a leaving group, poly(amino acid), protein, polysaccharide, or label.
12. The conjugate of claim 11 wherein X is (CH 2 ) 2 and Y is O.