Journal article
Affinity Purification of Antibodies Specific for 1,4-Dihydropyridine Ca2+ Channel Blockers
Circulation research, Vol.61(4 Suppl I), pp.I-37-I-45
10/1987
PMID: 3308157
Abstract
High-affinity antibodies specific for the 1,4-dihydropyridine Ca channel blockers have been produced in sheep and affinity purified using a dihydropyridine-Sepharose affinity column. Dihydropyridine-Sepharose affinity matrix was synthesized by reaction of amlnohexyl-Sepharose with an affinity analogue of nifedipine, dimethyl l,4-dihydro-2,6-dimethyl-4-(2-isothiocyanatophenyl)-3,5-pyridinedicarboxylate. Residual amine groups were then blocked by carbodiimide-catalyzed acetylation. [H]Nitrendiplne-binding activity in serum was specifically absorbed by the dihydropyridine-Sepharose affinity column. The bound antibody was eluted with diethylamine (pH 11.5) in 10percent; dioxane or with a low-affinity dihydropyridine ligand (diethyl 1,4-dihydro-2,4,6-trimethyl-3,5-pyridinedicarboxylate), pH 7.4. Thirty-six milligrams of highly pure IgG antibody, as demonstrated by sodium dodecyl sulfate-gel electrophoresis, was isolated from 50 ml hyperimmune sheep serum. The affinity-purified anti-dihydropyridine antibodies have been shown to have high affinity (Kd ~ 0.1 nM) and specificity for the 1,4-dihydropyridine Ca channel blockers and, therefore, exhibit dihydropyridine-binding properties similar to the membrane receptor for the 1,4-dihydropyridine Ca channel blockers. Immunoblot staining of an azidopine-bovine serum albumin conjugate with affinity-purified anti-dihydropyridine antibodies demonstrated that the anti-dihydropyridine antibodies recognize the 1,4-dihydropyridine Ca channel blockers when covalently coupled to protein and, therefore, should be useful in the identification and purification of receptors covalently labelled with 1,4-dihydropyridine Ca channel blockers.
Details
- Title: Subtitle
- Affinity Purification of Antibodies Specific for 1,4-Dihydropyridine Ca2+ Channel Blockers
- Creators
- Alan Sharp - From the Department of Physiology and Biophysics, The University of Iowa, Iowa City, IowaKevin Campbell
- Resource Type
- Journal article
- Publication Details
- Circulation research, Vol.61(4 Suppl I), pp.I-37-I-45
- Publisher
- American Heart Association, Inc
- PMID
- 3308157
- ISSN
- 0009-7330
- eISSN
- 1524-4571
- Language
- English
- Date published
- 10/1987
- Academic Unit
- Neurology; Molecular Physiology and Biophysics; Iowa Neuroscience Institute
- Record Identifier
- 9984068368002771
Metrics
33 Record Views