Compositions and Methods for Detecting Autoantibodies

Abstract

The invention provides compositions and methods for detecting thyroid hormone blocking immunoglobulin (TBI). The invention's methods are sensitive and specific for TBI, and may be used for the dual detection of both TBI and TSI. The invention's compositions and methods are useful for the diagnosis of diseases that are associated with the presence of TBI and/or TSI, for monitoring the progress of disease and/or treatment regimens, therapeutics, vaccines, etc., and for assisting clinicians in making treatment decisions.

1 - 20 . (canceled) 
     
     
         21 . A method for detecting thyroid hormone blocking immunoglobulin (TBI) in a biologic sample, comprising
 i) combining:
 a) transgenic cells stably transfected with 
 a first nucleic acid sequence which encodes a reporter wherein the first nucleic acid is operably linked to a cAMP-inducible promoter, and a second nucleic acid sequence which encodes a chimeric thyroid stimulating hormone receptor (TSHR), wherein the second nucleic acid sequence is operably linked to a constitutive promoter; and 
 b) a thyroid stimulating polypeptide that stimulates the chimeric TSHR upon binding to the chimeric TSHR, to generate a reagent mix; 
   ii) combining the reagent mix with
 a) a control sample to generate a first sample; or 
 b) a test sample comprising the biological sample to generate a second sample; and 
   iii) measuring the level of expression of said reporter in said first sample and in said second sample, wherein a reduced level of expression of said reporter in said second sample compared to said first sample indicates the presence of TBI in said biologic sample.   
     
     
         22 . The method of  claim 21 , further comprising determining the level of TBI in the biologic sample comprising measuring the level of expression of said reporter in one or more standard samples which each contain a known concentration of thyroid stimulating hormone (TSH) and comparing the level of expression of said reporter in said second sample with the level of expression of said reporter in said one or more standard samples. 
     
     
         23 . The method of  claim 21 , wherein said first nucleic acid sequence encodes a protein which can generate a fluorescent or a bioluminescent signal. 
     
     
         24 . The method of  claim 21 , wherein the first nucleic acid sequence encodes the amino acid sequence of SEQ ID NO:3. 
     
     
         25 . The method of  claim 21 , wherein the transgenic cells are Chinese hamster ovary (CHO) cells or human Rhabdomyosarcoma (RD) cells. 
     
     
         26 . The method of  claim 21 , wherein the thyroid stimulating polypeptide is selected from the group consisting of TSH, a thyroid stimulating monoclonal antibody, an a thyroid stimulating polyclonal antibody. 
     
     
         27 . The method of  claim 21 , wherein the constitutive promoter is an SV40 promoter sequence. 
     
     
         28 . The method of  claim 21 , wherein the second nucleic acid comprises SEQ ID NO:2. 
     
     
         29 . The method of  claim 21 , wherein the thyroid stimulating polypeptide is TSH and wherein said TSH is present in at least one of said first sample at a concentration of about 0.2 mIU/ml to 100 mIU/ml. 
     
     
         30 . The method of  claim 22 , wherein the IC 50  for TBI is about 5-fold to 15-fold smaller than the IC 50  for TBI when the method is performed with said transgenic cells stably transfected with wild type TSHR. 
     
     
         31 . The method of  claim 22 , wherein the IC 50  for TBI is about 10-fold to 30-fold smaller when the thyroid stimulating polypeptide is TSH than when the thyroid stimulating polypeptide is an anti-TBI monoclonal antibody. 
     
     
         32 . A kit comprising
 a) transgenic cells stably transfected with a first nucleic acid sequence which encodes a reporter wherein the first nucleic acid is operably linked to a cAMP-inducible promoter, and   a second nucleic acid sequence which encodes a chimeric thyroid stimulating hormone receptor (TSHR), wherein the second nucleic acid sequence is operably linked to a constitutive promoter, wherein said cells express a chimeric TSHR on the cell membrane;   b) a thyroid stimulating polypeptide that stimulates the chimeric TSHR upon binding to the chimeric TSHR on the cell membrane;   c) instructions for use.   
     
     
         33 . The kit of  claim 32 , wherein said kit further comprises a positive control sample that contains thyroid hormone blocking immunoglobulin (TBI). 
     
     
         34 . The kit of  claim 32 , wherein said thyroid stimulating polypeptide is thyroid stimulating hormone (TSH). 
     
     
         35 . The kit of  claim 32 , wherein said instructions are for detecting thyroid hormone stimulating immunoglobulin (TSI) in a biologic sample. 
     
     
         36 . The kit of  claim 32 , wherein said kit further comprises a thyroid hormone stimulating immunoglobulin (TSI).