Monoclonal antibody to a human thymocyte antigen and methods of preparing same
AssigneeOrtho Pharmaceutical Corporation
InventorsPatrick C. Kung, Gideon Goldstein
Life of this patent
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Abstract
Hybrid cell line for production of monoclonal antibody to an antigen found on approximately 70% of normal human thymocytes. The hybrid is formed by fusing splenocytes from immunized CAF.sub.1 mice with P3X63Ag8U1 myeloma cells. Diagnostic and therapeutic uses of the monoclonal antibody are also disclosed.
Claims11
A monoclonal antibody of class IgG produced by a hybridoma formed by fusion of cells from a mouse myeloma line and spleen cells from a mouse previously immunized with human thymocytes, which antibody:
- (a) reacts with approximately 70 percent of normal human thymocytes, but not with normal human peripheral T cells, B cells, Null cells, or bone marrow cells,
- (b) reacts with about 65 percent of untreated thymocytes, 10 percent of thymocytes previously treated with monoclonal antibody produced by hybrid cell line ATCC CRL 8002 and complement, 82 percent of thymocytes previously treated with monoclonal antibody produced by hybrid cell line ATCC CRL 8001 and complement, and 13 percent of the thymocytes previously treated with monoclonal antibody produced by hybrid cell line ATCC CRL 8014 and complement;
- (c) reacts with stage II T-cell ALL but not with stage I or stage III T-cell ALL;
- (d) reacts with CEM and MOLT-4 cell lines but not with HSB-2 cell line;
- and
- (e) defines a T cell population which is lower than normal levels in primary biliary cirrhosis, multiple sclerosis, and hyper IgE, higher than normal levels in acute graft versus host reaction, and completely absent in myasthenia gravis, acute infectious mononucleosis, all stages of Hodgkins disease, and psoriasis.
A monoclonal antibody of class IgG produced by a hybridoma formed by fusion of cells from a mouse myeloma line and spleen cells from a mouse previously immunized with human thymocytes which reacts with approximately 70 percent of normal human thymocytes, but not with normal human peripheral T cells, B cells, Null cells, or bone marrow cells.
Mouse monoclonal antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells.
A monoclonal antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells, prepared by the method which comprises the steps of:
- (i) immunizing mice with normal human thymocytes;
- (ii) removing the spleens from said mice and making a suspension of the spleen cells;
- (iii) fusing said spleen cells with mouse myeloma cells in the presence of a fusion promoter;
- (iv) diluting and culturing the fused cells in separate wells in a medium which will not support the unfused myeloma cells;
- (v) evaluating the supernatant in each well containing a hybridoma for the presence of antibody to E rosette positive purified T cells or human thymocytes;
- (vi) selecting and cloning a hybridoma producing antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells;
- and
- (vii) recovering the antibody from the supernatant above said clones.
A monoclonal antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells, prepared by the method which comprises the steps of:
- (i) immunizing mice with purified human thymocytes;
- (ii) removing the spleens from said mice and making a suspension of the spleen cells;
- (iii) fusing said spleen cells with mouse myeloma cells in the presence of a fusion promoter;
- (iv) diluting and culturing the fused cells in separate wells in a medium which will not support the unfused myeloma cells;
- (v) evaluating the supernatant in each well containing a hybridoma for the presence of antibody to E rosette positive purified T cells or human thymocytes;
- (vi) selecting and cloning a hybridoma producing antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells;
- (vii) transferring said clones intraperitoneally into mice;
- and
- (viii) harvesting the malignant ascites or serum from said mice, which ascites or serum contains the desired antibody.
A method of preparing monoclonal antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells, which comprises culturing the hybridoma ATCC CRL 8020 in a suitable medium and recovering the antibody from the supernatant above said hybridoma.
A method of preparing monoclonal antibody which reacts with approximately 70% of normal human thymocytes but not with normal human peripheral T cells, B cells, null cells, or bone marrow cells, which comprises injecting into a mouse the hybridoma ATCC CRL 8020 and recovering the antibody from the malignant ascites or serum of said mouse.
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