Methods of using chimeric receptors to identify autoimmune disease
View Patent ↗The present invention provides methods and compositions useful in the diagnosis and management of autoimmune diseases. In particular, the present invention provides improved methods and compositions for the diagnosis and management of Graves' disease. The methods of the present invention not only avoids the need for radioactivity and are much simpler, economical, and rapid than methods traditionally used for the diagnosis of Graves' disease, but also improve upon the sensitivity and detection abilities of previous luciferase-based autoantibody detection assays.
1. A method for detecting thyroid-stimulating antibodies, comprising:
a) combining:
1) a sample suspected of containing thyroid-stimulating auto-antibodies and
2) a recombinant cell line comprising a chimeric thyroid stimulating hormone (TSH) receptor gene and a reporter gene, wherein the chimeric TSH receptor gene encodes a polypeptide by nucleotides 45 to 2234 of SEQ ID NO:3, said polypeptide comprising, in an N-terminal to C-terminal direction, the N-terminal portion of the human thyroid stimulating hormone receptor (hTSHR), a portion of the rat leutinizing hormone (rLH) receptor wherein said rLH portion comprises the amino acid sequence encoded by nucleotides 828-1046 of SEQ ID NO:3, and the C-terminal portion of the hTSHR;
b) incubating the sample and recombinant cell line under conditions such that the reporter gene generates and detectable signal; and
c) detecting the signal generated by the reporter gene.
2. The method of claim 1 , wherein the sample is a serum sample.
3. The method of claim 1 , wherein the reporter gene is selected from the group consisting of a bacterial β-galactosidase gene, a bacterial chloramphenicol acetyltransferase gene, a luciferase gene, and a β-glucuronidase gene.
4. The method of claim 1 , wherein the reporter gene is a firefly luciferase gene.
5. The method of claim 1 , wherein the reporter gene allows detection of cAMP.
6. The method of claim 1 , wherein the reporter gene is operably linked to a glycoprotein hormone alpha subunit promoter.
7. The method of claim 1 , wherein the method further comprising measuring the intensity of the signal, wherein the intensity positively correlates with thyrotropin stimulating hormone receptor auto antibody concentration present in the sample.
8. The method of claim 1 , wherein the detecting comprising using a luminometer.
9. The method of claim 1 , wherein the recombinant cell line is an RD cell line or a CHO cell line.
10. The method of claim 1 , wherein the recombinant cell line is stably transfected with the reporter gene.
11. The method of claim 1 , wherein the recombinant cell line is stably transfected with the chimeric TSH receptor gene.
12. The method of claim 1 , wherein the chimeric TSH receptor gene comprises SEQ ID NO:3.