Monoclonal antibodies to the thyrotropin receptor raised by an autoantiidiotypic protocol and their relationship to monoclonal autoantibodies from Graves' patients
Monoclonal antibodies that bind to the TSH receptor were obtained by an autoantiidiotypic approach in which immunization of BALB/c mice was performed with mixtures of bovine (b) and human (h) TSH. Two of 28 positive wells were selected for cloning and characterization: D2 and 4G11. Their antiidiotypic character was evidenced by TSH-inhibitable binding to affinity-purified polyclonal anti-TSH. The specificity of D2 and 4G11 for the hormone-binding region of the TSH receptor was demonstrated by several findings: 1) they inhibited the binding of (125I)iodo-bTSH to receptor in a dose-dependent manner; 2) their binding to partially purified thyroid plasma membranes could be completely inhibited by bTSH and hTSH; and 3) they inhibited the TSH-dependent growth and adenylate cyclase stimulation in FRTL-5 cells in a dose-dependent manner. By Western blot analysis of bovine thyroid membranes, D2 bound to a polypeptide of 188,000-195,000 mol wt under nonreducing conditions and 54,000-59,000 mol wt after treatment of membranes with beta-mercaptoethanol; the 4G11 epitope was undetectable. Scatchard analysis of the binding of 125I-labeled antibodies to receptor showed that 4G11 bound to a single site with a Kd of 5.7 X 10(-9) M, whereas D2 showed complex binding characterized by high affinity (Kd = 1.74 X 10(-11) M) and low affinity (Kd = 1.3 X 10(-8) M) sites. Binding studies in which D2 and 4G11 competed with each other for the TSH receptor showed mutual but unequal inhibition. The data suggest that portions of the D2 and 4G11 epitopes overlap, but that there is a high affinity binding site(s) for D2 for which 4G11 competes less effectively. The binding of D2 and 4G11 to TSH receptor was inhibited by monoclonal antibodies secreted by Graves' heterohybridomas, showing that D2 and 4G11 share characteristics with autoantibodies of Graves' disease.
- Research Organization:
- Columbia Univ., New York, NY (USA)
- OSTI ID:
- 6975105
- Journal Information:
- Endocrinology; (United States), Vol. 122:6
- Country of Publication:
- United States
- Language:
- English
Similar Records
Microheterogeneous forms of radioiodinated bovine thyrotropin: discrimination of different receptor-active components by gel permeation chromatography
Binding of bovine thyrotropin to specific sites in thyroid tissue from control and hemithyroidectomized rats
Related Subjects
ENDOCRINE DISEASES
PATHOGENESIS
MONOCLONAL ANTIBODIES
BIOCHEMICAL REACTION KINETICS
SPECIFICITY
TSH
RECEPTORS
AFFINITY
CATTLE
CELL MEMBRANES
CYCLASES
DOSE-RESPONSE RELATIONSHIPS
HYBRIDOMAS
IMMUNOASSAY
IODINE 125
MICE
MOLECULAR WEIGHT
PATIENTS
THYROID
TRACER TECHNIQUES
ANIMAL CELLS
ANIMALS
ANTIBODIES
BETA DECAY RADIOISOTOPES
BODY
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
DISEASES
DOMESTIC ANIMALS
ELECTRON CAPTURE RADIOISOTOPES
ENDOCRINE GLANDS
ENZYMES
GLANDS
HORMONES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LYASES
MAMMALS
MEMBRANE PROTEINS
MEMBRANES
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDE HORMONES
PITUITARY HORMONES
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RODENTS
RUMINANTS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques
550901 - Pathology- Tracer Techniques