Expression analysis of the thyrotropin‐releasing hormone receptor (TRHR) in the immune system using agonist anti‐TRHR monoclonal antibodies

FEBS Letters

García San Frutos, Miriam²  Mellado, Mario³  Rodrı́guez-Frade, José Miguel³  Martı́nez-A, Carlos³  de la Peña, Pilar¹  Fernández-Agulló, Teresa²  Montoya, Eladio² 

[1]Departamento de Biologı́a Funcional, Area de Bioquı́mica, Facultad de Medicina, Universidad de Oviedo, E-33006 Oviedo, Spain

[2]Departamento de Fisiologı́a, Facultad de Medicina, Universidad de Alcalá de Henares, E-28871 Madrid, Spain

[3]Department of Immunology and Oncology, Centro Nacional de Biotecnologı́a (CSIC), Campus de Cantoblanco, E-28049 Madrid, Spain

关键词: Thyrotropin-releasing hormone;    Receptor;    Monoclonal antibody;    Immune system;    EIA;    enzyme-linked immunoassay;    GAM;    goat anti-mouse immunoglobulin antibody;    FITC;    fluorescein isothiocyanate;    KLH;    keyhole limpet hemocyanin;    PBMC;    peripheral blood mononuclear cells;    PE;    phycoerythrin;    PHA;    phytohemagglutinin;    PO;    peroxidase;    SF;    supernatant fluid;    TRHR;    yhyrotropin-releasing hormone receptor;    TBS;    Tris-buffered saline;   

DOI  :  10.1016/S0014-5793(99)00607-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Monoclonal anti-rat thyrotropin-releasing hormone (TRH) receptor (TRHR)-specific antibodies (mAb) were generated by immunization with synthetic peptides of rat TRHR partial amino acid sequences; one (TRHR01) was directed against a sequence (84–98) in the extracellular portion of the rat TRHR reported to be constant among different species, including man, and the second (TRHR02) recognizes the C-terminal region sequence 399–412. In lysates from GH₄C₁ cells, a clonal rat pituitary cell line, both mAb recognize the TRHR in Western blot analysis, and TRHR02 immunoprecipitates the TRHR. Incubation of GH₄C₁ cells with the mAb causes a fluorescence shift in fluorescence-activated cell sorting analysis. The cells were stained specifically by both mAb using immunocytochemical techniques. Furthermore, TRHR01 is agonistic in its ability to trigger Ca²⁺ flux, and desensitizes the TRH receptor. We tested for TRHR in several rat organs and found expression in lymphoid tissues. TRHR01 recognizes the human TRHR, and analysis of human peripheral blood lymphocyte and tonsil-derived leukocyte populations showed receptor expression in non-activated and phytohemagglutinin-activated T and B cells.

【 授权许可】

Unknown