EP0235113A2
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Immortalization of primary cells
Assignee: SmithKline Beecham Corp
•
Filing: 1987-02-26
•
Granted: —
•
Expires: 2007-02-26
Abstract
Abstract The use of electroporation as a transfection method to immortalize primary cells with oncogenic DNA/oncogene(s).
Inventors
- 👤 Edward Mark Henri
- 👤 Zdenka Ludmila Jonak
- 👤 Patrick Simon Jacques Machy
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-06 05:34
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/87; C07; C07K; C07K14/00; C07K14/435; C07K14/46; C07K14/47; C07K14/4701; C07K14/4746; C07K14/82; C12N15/88
Claims Analysis
Total Claims: 86
Claim 1
Independent Claim
52 words
A method for producing an immortalized primary cell line which comprises: (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic DNA/oncogene (s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; and (c) recovering from such transfected primary cells an immortalized primary cell line.
Claim 2
Independent Claim
52 words
A method for producing an immortalized primary cell line which comprises: (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic DNA/oncogene (s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; and (c) recovering from such transfected primary cells an immortalized primary cell line.
Claim 3
Dependent Claim
41 words
The method of Claim 1 wherein the oncogenic D NA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be immortalized.
Claim 4
Dependent Claim
39 words
The method of Claim 1 wherein the oncogene(s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immortalized.
Claim 5
Dependent Claim
13 words
The method of Claim 2 wherein the primary cells are of hematopoietic origin.
Claim 6
Dependent Claim
30 words
The method of Claim 3 wherein the primary cells are multipotential stem cells, B-lymphocytes, 3-lineage precursors, T-lympocytes, T-lineage precursors, nyeloid stem cells, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes or mast cells.
Claim 7
Dependent Claim
11 words
The method of Claim 4 wherein the primary cells are B-lymphocytes.
Claim 8
Dependent Claim
15 words
The method of Claim 5 wherein the B lymphocytes have been activated/stimulated with an antigen.
Claim 9
Dependent Claim
17 words
The method of Claim 5 wherein the oncogene (s) is the Ha-ras oncogene or the p53-H7 oncogene.
Claim 10
Dependent Claim
14 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from BJAB.
Claim 11
Dependent Claim
17 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 12
Dependent Claim
17 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 13
Dependent Claim
12 words
An immortalized primary cell line produced by the method of Claim 1.
Claim 14
Dependent Claim
41 words
The cell line of Claim 12 wherein the oncogenic DNA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be immortalized.
Claim 15
Dependent Claim
42 words
The cell line of Claim 12 wherein the oncogene (s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immor talized.
Claim 16
Dependent Claim
14 words
The cell line of Claim 12 wherein the primary cells are of hematopoietic origin.
Claim 17
Dependent Claim
32 words
The cell line of Claim 15 wherein the primary cells are multipotential stem cells, B -lymphocytes, B-lineage precursors, T-lympocytes, T-lineage precursors, myeloid stem cells, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes or mast cells.
Claim 18
Dependent Claim
12 words
The cell line of Claim 16 wherein the primary cells are B-lymphocytes.
Claim 19
Dependent Claim
16 words
The cell line of Claim 17 wherein the B lymphocytes have been activated/stimulated with an antigen.
Claim 20
Dependent Claim
13 words
The cell line of Claim 16 wherein the oncogene(s) is the Ha-ras oncogene.
Claim 21
Dependent Claim
14 words
The cell line of Claim 16 wherein oncogenic DNA is genomic DNA from BJAB.
Claim 22
Dependent Claim
18 words
The cell line of Claim 16 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 23
Dependent Claim
18 words
The cell line of Claim 16 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 24
Independent Claim
80 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within an immortalized primary cell line which comprises: (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic DNA/oncogene (s) (which, optionally is comprised by liposomes) by electroporation thereby producing transfected primary cells; (c) recovering from such transfected primary cells the immortalized primary cell line; and (d) culturing such immortalized cell line under conditions which enable the production of such biological products.
Claim 25
Independent Claim
80 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within an immortalized primary cell line which comprises: (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic DNA/oncogene (s) (which, optionally is comprised by liposomes) by electroporation thereby producing transfected primary cells; (c) recovering from such transfected primary cells the immortalized primary cell line; and (d) culturing such immortalized cell line under conditions which enable the production of such biological products.
Claim 26
Independent Claim
71 words
A method for production of a contransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises: (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and the foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; and (c) recovering from such cotransfected primary cells an immortalized primary cell line containing the foreign functional DNA sequence;
Claim 27
Independent Claim
71 words
A method for production of a contransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises: (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and the foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; and (c) recovering from such cotransfected primary cells an immortalized primary cell line containing the foreign functional DNA sequence;
Claim 28
Dependent Claim
41 words
The method of Claim 24 wherein the oncogenic DNA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be i immortalized.
Claim 29
Dependent Claim
40 words
The method of Claim 24 wherein the oncogene(s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immor talized.
Claim 30
Dependent Claim
13 words
The method of Claim 24 wherein the primary cells are of hematopoietic origin.
Claim 31
Dependent Claim
30 words
The method of Claim 27 wherein the primary cells are multipotential stem cells, B-lymphocytes, B-lineage precursors, T-lympocytes, T-lineage precursors, myeloid stem cells, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes or mast cells.
Claim 32
Dependent Claim
11 words
The method of Claim 28 wherein the primary cells are B-lymphocytes.
Claim 33
Dependent Claim
15 words
The method of Claim 29 wherein the B lymphocytes have been activated/stimulated with an antigen.
Claim 34
Dependent Claim
17 words
The method of Claim 29 wherein the oncogene (s) is the Ha-ras oncogene or the p53-H7 oncogene.
Claim 35
Dependent Claim
14 words
The method of Claim 29 wherein the oncogenic DNA is genomic DNA from BJAB.
Claim 36
Dependent Claim
17 words
The method of Claim 29 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 37
Dependent Claim
17 words
The method of Claim 29 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 38
Dependent Claim
13 words
A cotransfected immortalized primary cell line produced by the method of Claim 24.
Claim 39
Dependent Claim
43 words
36 . The cell line of Claim 35 wherein the oncogenic DNA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be immortalized.
Claim 40
Dependent Claim
41 words
The cell line of Claim 35 wherein the oncogene (s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immortalized.
Claim 41
Dependent Claim
14 words
The cell line of Claim 35 wherein the primary cells are of hematopoietic origin.
Claim 42
Dependent Claim
31 words
The cell line of Claim 38 wherein the primary cells are multipotential stem cells, B-lymphocytes, B-lineage precursors, T-lymphocytes, T-lineage precursors, myeloid stem cells, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes or mast cells.
Claim 43
Dependent Claim
12 words
The cell line of Claim 39 wherein the primary cells are B-lymphocytes.
Claim 44
Dependent Claim
15 words
The cell line'of Claim 40 wherein the B lymphocytes have been activated/stimulated with an antigen.
Claim 45
Dependent Claim
13 words
The cell line of Claim 40 wherein the oncogene(s) is the Ha-ras oncogene.
Claim 46
Dependent Claim
15 words
The cell line of Claim 40 wherein the oncogenic DNA is genomic DNA from BJAB.
Claim 47
Dependent Claim
18 words
The cell line of Claim 40 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 48
Dependent Claim
18 words
The cell line of Claim 40 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 49
Independent Claim
100 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises: (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and a foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; (c) recovering from such cotransfected primary cells the cotransfected immortalized primary cell line containing the foreign functional DNA sequence; and (d) culturing such cotransfected immortalized cell line under conditions which enable the production of such biological products.
Claim 50
Independent Claim
100 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises: (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and a foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; (c) recovering from such cotransfected primary cells the cotransfected immortalized primary cell line containing the foreign functional DNA sequence; and (d) culturing such cotransfected immortalized cell line under conditions which enable the production of such biological products.
Claim 51
Dependent Claim
25 words
An immortalized primary cell line provided that such immortalization is the result of transfection of a normal primary cell with DNA consisting essentially of oncogene(s).
Claim 52
Dependent Claim
25 words
An immortalized primary cell line provided that such immortalization is the result of transfection of a normal primary cell with DNA consisting essentially of oncogene(s).
Claim 53
Dependent Claim
38 words
The cell line of Claim 47 wherein the oncogene(s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type immortalized.
Claim 54
Dependent Claim
14 words
The cell line of Claim 47 wherein the primary cell is of hematopoietic origin.
Claim 55
Dependent Claim
32 words
The cell line of Claim 49 wherein the primary cell is a multipotential stem cell, B-lymphocyte, B-lineage precursor, T-lympocyte, T-lineage precursor, myeloid stem cell, monocyte, macrophage, neutrophil, eosinophil, megakaryocyte or mast cell.
Claim 56
Dependent Claim
13 words
The cell line of Claim 50 wherein the primary cell is a B-lymphocyte.
Claim 57
Dependent Claim
17 words
The cell line of Claim 51 wherein the oncogene(s) is the Ha-ras oncogene or the p53-H7 oncogene.
Claim 58
Dependent Claim
9 words
Claims for the Contractina States : AT. ES, GR
Claim 59
Independent Claim
53 words
A method for producing an immortalized primary cell line which comprises : (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic D NA/oncogene(s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; and (c) recovering from such transfected primary cells an immortalized primary cell line.
Claim 60
Independent Claim
53 words
A method for producing an immortalized primary cell line which comprises : (a) providing normal, activated/stimulated primary cells; (b) transfecting such primary cells with oncogenic D NA/oncogene(s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; and (c) recovering from such transfected primary cells an immortalized primary cell line.
Claim 61
Dependent Claim
42 words
2 . The method of. Claim 1 wherein the oncogenic DNA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be immortalized.
Claim 62
Dependent Claim
39 words
The method of Claim 1 wherein the oncogene(s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immortalized.
Claim 63
Dependent Claim
13 words
The method of Claim 2 wherein the primary cells are of hematopoietic origin.
Claim 64
Dependent Claim
30 words
The method of Claim 3 wherein the primary cells are multipotential stem cells, B-lymphocytes, B-lineage precursors, T-lymphocytes, T-lineage precursors, myeloid stem cells, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes or mast cells.
Claim 65
Dependent Claim
11 words
The method of Claim 4 wherein the primary cells are B-lymphocytes.
Claim 66
Dependent Claim
14 words
The method of Claim 5 wherein the B-lymphocytes have been activated/stimulated with an antigen.
Claim 67
Dependent Claim
16 words
The method of Claim 5 wherein the oncogene(s) is the Ha-ras oncogene or the p53-H7 oncogene.
Claim 68
Dependent Claim
14 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from BJAB.
Claim 69
Dependent Claim
17 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 70
Dependent Claim
17 words
The method of Claim 6 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 71
Independent Claim
80 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within an immortalized primary cell line which comprises : (a) providing normal, activated/stimulated primary cells. (b) transfecting such primary cells with oncogenic DNA/oncogene(s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; (c) recovering from such transfected primary cells the immortalized primary cell line; and (d) culturing such immortalized cell line under conditions which enable the production of such biological products.
Claim 72
Independent Claim
80 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within an immortalized primary cell line which comprises : (a) providing normal, activated/stimulated primary cells. (b) transfecting such primary cells with oncogenic DNA/oncogene(s) (which, optionally, is comprised by liposomes) by electroporation thereby producing transfected primary cells; (c) recovering from such transfected primary cells the immortalized primary cell line; and (d) culturing such immortalized cell line under conditions which enable the production of such biological products.
Claim 73
Independent Claim
72 words
A method for production of a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises : (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and the foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells, and (c) recovering from such cotransfected primary cells an immortalized primary cell line containing the foreign functional DNA sequence;
Claim 74
Independent Claim
72 words
A method for production of a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises : (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and the foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells, and (c) recovering from such cotransfected primary cells an immortalized primary cell line containing the foreign functional DNA sequence;
Claim 75
Dependent Claim
40 words
The method of Claim 13 wherein the oncogenic DNA is genomic DNA from a primary cell tumor or a tumor primary-cell line wherein such tumor or tumor cell-line has an origin corresponding to the primary cell type to be immortalized.
Claim 76
Dependent Claim
39 words
The method of Claim 13 wherein the oncogene(s) is/are from a primary cell tumor or a tumor primary cell line wherein such tumor or tumor cell line has an origin corresponding to the primary cell type to be immortalized.
Claim 77
Dependent Claim
15 words
1 6. The method of Claim 13 wherein the primary :ells are of hematopoietic origin.
Claim 78
Dependent Claim
32 words
17 . The method of Claim 16 wherein the primary :ells are multipotential stem cells, B-lymphocytes, B-Lineage precursors. T-lymphocytes, T-lineage precursors, myeloid stem cells, monocytes, macrophages, neutrophils. eosinophils, megakaryocytes or mast cells.
Claim 79
Dependent Claim
11 words
The method of Claim 17 wherein the primary cells are B-lymphocytes.
Claim 80
Dependent Claim
17 words
1 9. The method of Claim 18 wherein the B lymphocytes have been activated/stimulated with an antigen.
Claim 81
Dependent Claim
18 words
2 0. The method of Claim 18 wherein the oncogene(s) is the Ha-ras oncogene or the p53-H7 oncogene.
Claim 82
Dependent Claim
14 words
The method of Claim 18 wherein the oncogenic DNA is genomic DNA from BJAB.
Claim 83
Dependent Claim
19 words
2 2. The method of Claim 18 wherein the oncogenic DNA is genomic DNA from the REH cell line.
Claim 84
Dependent Claim
19 words
2 3. The method of Claim 18 wherein the oncogenic DNA is genomic DNA from the EL4 cell line.
Claim 85
Independent Claim
101 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises : (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and a foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; (c) recovering from such cotransfected primary cells the cotransfected immortalized primary cell line containing the foreign functional DNA sequence; and (d) culturing such cotransfected immortalized cell line under conditions which enable the production of such biological products.
Claim 86
Independent Claim
101 words
A method of producing biological products whose coding sequence is comprised by the DNA contained within a cotransfected immortalized primary cell line containing a foreign functional DNA sequence which comprises : (a) providing normal, activated/stimulated primary cells; (b) cotransfecting such primary cells with oncogenic DNA/oncogene(s) and a foreign functional DNA sequence (which, optionally, are comprised by liposomes) by electroporation thereby producing cotransfected primary cells; (c) recovering from such cotransfected primary cells the cotransfected immortalized primary cell line containing the foreign functional DNA sequence; and (d) culturing such cotransfected immortalized cell line under conditions which enable the production of such biological products.
References Cited (Backward Citations)
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