| Background and objective: PAR-1 clone was identified that encoded a protein of 425 residues with 7 hydrophobic domains of a typical GPCR. It is found in a wide variety of cell types, including platelets, endothelial cells, fibroblasts, monocytes, T cell lines, osteoblastlike cells, smooth muscle cells, neurons cells, glial ceils and in certain tumor cell lines. PAR-1 participated in the generation and development of all kinds of diseases, is involved in many physiological functions. Therefore, it is meaningful to prepare and identify monoclonal antibodies (mAbs) against PAR-1.Methods: BALB/c mice were immunized with 13 amino acid peptide of PAR-1 and hybridoma cell lines were obtained by fusing mouse spleen cells with myeloma NS-1 cells. The specificity of mAbs were characterized by enzyme-linked immunosorbent assay (ELISA), dot blot, immunohistochemical staining, flow cytometry (FCM) and confocal laser scanning microscopy (CLSM).Results: 2 hybridoma cell lines that secreted the mAbs against PAR-1 were obtained. One mAb was IgM, and the other one was IgG2b. Immunohistochemical staining revealed that the reactivities of mAbs to the lymphocyte cells in tonsil and colon tissues, to fibroblast cells in foreskin tissues, to macrophage in lung tissues were observed. FCM showed that the 2 mAbs reacted to PAR-1 expressed both on the membrane surface and in the cytoplasm of A549 cells. CLSM examination showed that the fluorescent markers were located both on the membrane and in the cytoplasm of A549 cells.Conclusion: The mAbs against PAR-1 are prepared successfully, which provides valuable tool for studies on inflammatory diseases.
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