Monoclonal-antibody-producing hybrid cell line, antibody and method of preparing it, therapeutic composition containing it and its diagnostic and therapeutic uses

Claim 1

A monoclonal antibody of class IgG produced by a hybridoma formed by fusion of spleen cells from a mouse previously immunized with human T cells and cells from a mouse myeloma line, which antibody: a) reacts with essentially all normal human peripheral T cells and cutaneous T lymphoma cells, but not with normal human peripheral B cells, null cells or macrophages; b) reacts with from about 5% to about 10% of normal human thymocytes; c) reacts with leukemic cells from humans with T cell chronic lymphoblastic leukemia but does not react with leukemic cells from humans with T cell acute lymphoblastic leukemia, null cell acute lymphoblastic leukemia, or B cell chronic lymphatic leukemia; d) reacts weakly with the human T cell line HJD-1 but does not react with CEM, Laz 191, or HMl; e) does not react with the Epstein-Barr virus- transformed human B cell lines Laz 007, Laz 156, Laz 256, or SB; and f) fixes complement.

Claim 18

A method for determining in an individual the proportion of circulating lymphocytes that are T cells which comprises mixing the antibody any one of Claims 1 to 4 with a circulating lymphocyte composition from said individual and determining the proportion of the circulating lymphocytes which react with said antibody, and are thus T cells.

Claim 19

A method for determining in an individual the proportion of circulating lymphocytes that are T cells which comprises mixing antibody produced from a hybridoma having the identifying characteristics of OKT3 with a circulating lymphocyte composition from said : individual and determining the proportion of the circulating lymphocytes which react with said antibody, and are thus T cells.

Claim 20

A method for determining in an individual the proportion of circulating lymphocytes that are T cells which comprises mixing antibody produced from a hybridoma having the identifying characteristics of OKT3 with a circulating lymphocyte composition from said : individual and determining the proportion of the circulating lymphocytes which react with said antibody, and are thus T cells.

Claim 21

A method for preparing monoclonal antibody which: a) reacts with essentially all normal human peripheral T cells and cutaneous T lymphoma cells, but not with normal human peripheral B cells, null cells or macrophages; b) reacts with from about 5% to about 10% of normal human thymocytes; c) reacts with leukemic cells from humans with T cell chronic lymphoblastic leukemia but does not react with leukemic cells from humans with T cell acute lymphoblastic leukemia, null cell acute lymphoblastic leukemia,'or B cell chronic lymphatic leukemia; d) reacts weakly with the human T cell line HJD-1 but does not react with CEM, Laz 191, or HM1; e) does not react with the Epstein-Barr virus- transformed human B cell lines Laz 007, Laz 156, Laz 256, or SB; and f) fixes complement, which comprises the steps of: i) immunizing mice with E rosette positive purified human T cells; ii) removing the spleens from said mice and making a suspension of 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 the desired antibody; vi) selecting and cloning hybridomas producing the desired antibody; and vii) recovering the antibody from the supernatant above said clones.

Claim 22

A method for preparing monoclonal antibody which: a) reacts with essentially all normal human peripheral T cells and cutaneous T lymphoma cells, but not with normal human peripheral B cells, null cells or macrophages; b) reacts with from about 5% to about 10% of normal human thymocytes; c) reacts with leukemic cells from humans with T cell chronic lymphoblastic leukemia but does not react with leukemic cells from humans with T cell acute lymphoblastic leukemia, null cell acute lymphoblastic leukemia,'or B cell chronic lymphatic leukemia; d) reacts weakly with the human T cell line HJD-1 but does not react with CEM, Laz 191, or HM1; e) does not react with the Epstein-Barr virus- transformed human B cell lines Laz 007, Laz 156, Laz 256, or SB; and f) fixes complement, which comprises the steps of: i) immunizing mice with E rosette positive purified human T cells; ii) removing the spleens from said mice and making a suspension of 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 the desired antibody; vi) selecting and cloning hybridomas producing the desired antibody; and vii) recovering the antibody from the supernatant above said clones.

Claim 24

A method for preparing monoclonal antibody which: a) reacts with essentially all normal human peripheral T cells and cutaneous T lymphoma cells, but not with normal human peripheral B cells, null cells or macrophages; b) reacts with from about 5% to about 10% of normal human thymocytes; c) reacts with leukemic cells from humans with T cell chronic lymphoblastic leukemia but does not react with leukemic cells from humans with T cell acute lymphoblastic leukemia, null cell acute lymphoblastic leukemia,.or B cell chronic lymphatic leukemia; d) reacts weakly with the human T cell line HJD-1 but does not react with CEM, Laz 191, or HM1; e) does not react with the Epstein-Barr virus- transformed human B cell lines Laz 007, Laz 156, Laz 256, or SB; and f) fixes complement, which comprises the steps of: i) immunizing mice with E rosette positive purified human T cells; 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 the desired antibody; vi) selecting and cloning hybridomas producing the desired antibody; vii) recovering the antibody from the supernatant above said clones; viii) transferring said clones intraperitoneally into mice; and ix) harvesting the malignant ascites or serum from said mice.

Claim 25

A method for preparing monoclonal antibody which: a) reacts with essentially all normal human peripheral T cells and cutaneous T lymphoma cells, but not with normal human peripheral B cells, null cells or macrophages; b) reacts with from about 5% to about 10% of normal human thymocytes; c) reacts with leukemic cells from humans with T cell chronic lymphoblastic leukemia but does not react with leukemic cells from humans with T cell acute lymphoblastic leukemia, null cell acute lymphoblastic leukemia,.or B cell chronic lymphatic leukemia; d) reacts weakly with the human T cell line HJD-1 but does not react with CEM, Laz 191, or HM1; e) does not react with the Epstein-Barr virus- transformed human B cell lines Laz 007, Laz 156, Laz 256, or SB; and f) fixes complement, which comprises the steps of: i) immunizing mice with E rosette positive purified human T cells; 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 the desired antibody; vi) selecting and cloning hybridomas producing the desired antibody; vii) recovering the antibody from the supernatant above said clones; viii) transferring said clones intraperitoneally into mice; and ix) harvesting the malignant ascites or serum from said mice.