title: Molecular dissection of an hCG-beta epitope using single-step solid phase radioimmunoassay.
abstract: BACKGROUND
  Peptides and proteins have both sequence-specific (contiguous) and conformation-specific (discontiguous) epitopes. Sequence-specific epitopes are delineated by peptide approach and other robust methods like competition assays, gene expression assays, synthetic peptide library based assays etc. Available methods for delineation of conformation-specific epitopes are cumbersome (X-ray crystallography etc.), time-consuming and require costly sophisticated equipments. Hence, there is a need to develop a simple method for identification and mapping of conformation-specific epitopes.
  
  
  METHODS
  In the single-step solid phase radioimmunoassay (SS-SPRIA), an immunochemical bridge of 'mouse IgG-anti-mouse IgG' was prepared in the polypropylene wells followed by adsorption with hCG specific monoclonal antibody (MAb) G(1)G(10).1. The extent of competitive inhibition in binding ability of (125)IhCG-beta with chemically or enzymatically modified hCG-beta to immobilized MAb G(1)G(10).1 in comparison to hCG-beta standards was utilized to identify the epitopic amino acid involved in epitope-paratope interaction.
  
  
  RESULTS
  Data clearly suggest that the epitope under investigation consisted of Arg (94, 95) and Asp (99) at the core region with a Lys (104) and a His (106) in the proximity and absence of chymotrypsin susceptible Phe or Tyr in this region.
  
  
  CONCLUSIONS
  The data of SS-SPRIA revealed the 93-100 loop of amino acid sequence, as the core region of conformation-specific epitope of hCG-beta at or near the receptor-binding region. Hence, SS-SPRIA seems to be a simple method for identification and mapping of conformation-specific epitopes.
authors.affiliation: Department of Reproductive Biomedicine, National Institute of Health and Family Welfare, Munirka, New Delhi-110067, India.
authors.forename: Pramod V
authors.initials: PV
authors.lastname: Prasad
authors.forename: Shail K
authors.initials: SK
authors.lastname: Chaube
authors.forename: Manoj
authors.initials: M
authors.lastname: Panchal
authors.forename: Rajesh
authors.initials: R
authors.lastname: Chaudhary
authors.forename: K
authors.initials: K
authors.lastname: Muralidhar
authors.forename: Vishwajeet
authors.initials: V
authors.lastname: Rohil
authors.forename: G L
authors.initials: GL
authors.lastname: Kumari
authors.forename: Anand
authors.initials: A
authors.lastname: Kumar
authors.forename: B
authors.initials: B
authors.lastname: Ashish
authors.forename: G S
authors.initials: GS
authors.lastname: Murthy
authors.forename: T G
authors.initials: TG
authors.lastname: Shrivastav
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D018997:Chorionic Gonadotropin, beta Subunit, Human / Q000737:chemistry* / Q000276:immunology*; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D011863:Radioimmunoassay / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D018997:Chorionic Gonadotropin, beta Subunit, Human; D000939:Epitopes