Patent US4464465A successfully refreshed from Google Patents.
US4464465A
Expired / Lapsed
US
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Cell-driven viral transfer in eukaryotes
Assignee: Genetic Systems Corp
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Filing: 1982-04-05
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Granted: 1984-08-07
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Expires: 2002-04-05
Abstract
Abstract Novel method, cells and compositions are provided involving transforming B-lymphocytes to provide immortalization for continuous production of monoclonal antibodies to a predetermined ligand. T-cell free B-lymphocytes are combined with an Epstein-Barr virus transformed cell sensitive to a cytotoxic agent which does not significantly affect the B-lymphocytes under conditions where the sensitive EBV transformed cell acting as the transfer agent is killed and efficiently transforms the B-lymphocyte recipient cells with EBV. The EBV transformed B-lymphocyte cells are amplified and cloned, the desired clones isolated in accordance with conventional techniques and then used for production of monoclonal antibodies.
Inventors
- 👤 Mark E. Lostrom
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 20:01
CPC Classifications
C; C07; C07K; C07K16/00; Y; Y10; Y10S; Y10S435/00; Y10S435/8215; Y10S435/948
Claims Analysis
Total Claims: 22
Claim 1
Independent Claim
80 words
A method for immortalizing recipient immunoglobulin producing cells employing an EBV transformed cell as the transferring cell, said method comprising: co-cultivating said transferring and said recipient cells in the substantial absence of functional T-cells, for sufficient incubation times and conditions to transfer EBV to said recipient cells to produce immortalized Ig producing cells which are EBNA-positive and which have a normal diploid number of chromosomes and cloning said immortalized Ig producing cells in the virtual absence of said transferring cells.
Claim 2
Independent Claim
80 words
A method for immortalizing recipient immunoglobulin producing cells employing an EBV transformed cell as the transferring cell, said method comprising: co-cultivating said transferring and said recipient cells in the substantial absence of functional T-cells, for sufficient incubation times and conditions to transfer EBV to said recipient cells to produce immortalized Ig producing cells which are EBNA-positive and which have a normal diploid number of chromosomes and cloning said immortalized Ig producing cells in the virtual absence of said transferring cells.
Claim 3
Dependent Claim
28 words
A method according to claim 1, wherein at least one of said cocultivating and cloning is in the presence of a cytotoxic agent selective for said transferring cells.
Claim 4
Dependent Claim
20 words
A method according to claim 2, wherein said transferring cells are HAT sensitive and said cytotoxic selective agent is HAT.
Claim 5
Dependent Claim
19 words
A method according to claim 1, wherein said recipient cells are primed with a ligand prior to said cocultivation.
Claim 6
Dependent Claim
23 words
A method according to claim 1, wherein said recipient cells are from a host having a disease which stimulates a B-cell immune response.
Claim 7
Independent Claim
122 words
A method for producing immunoglobulins ("Ig") specific for a predetermined determinant site, said method comprising: co-cultivating under selectively cytotoxic conditions EBV-transformed cells sensitive to said selective conditions, as transferring cells, and B-lymphocytes activated by an immunogen containing said determinant site, as recipient cells, for sufficient incubation times and conditions, resulting in the transformation of said recipient cells to produce immortalized Ig producing cells which are EBNA-positive and which have a normal diploid number of chromosomes, and the mortality of said sensitive cells; expanding the resulting population of immortalized Ig producing cells in an appropriate nutrient medium in individual wells and selecting for production of the desired immunoglobulin from the resulting clones; and harvesting the desired immunoglobulin from supernatants from the desired clones.
Claim 8
Independent Claim
122 words
A method for producing immunoglobulins ("Ig") specific for a predetermined determinant site, said method comprising: co-cultivating under selectively cytotoxic conditions EBV-transformed cells sensitive to said selective conditions, as transferring cells, and B-lymphocytes activated by an immunogen containing said determinant site, as recipient cells, for sufficient incubation times and conditions, resulting in the transformation of said recipient cells to produce immortalized Ig producing cells which are EBNA-positive and which have a normal diploid number of chromosomes, and the mortality of said sensitive cells; expanding the resulting population of immortalized Ig producing cells in an appropriate nutrient medium in individual wells and selecting for production of the desired immunoglobulin from the resulting clones; and harvesting the desired immunoglobulin from supernatants from the desired clones.
Claim 9
Dependent Claim
13 words
A method according to claim 6, wherein said selective conditions are HAT medium.
Claim 10
Independent Claim
43 words
A method according to any one of claims 6 or 7, wherein said clones producing said desired immunoglobulin are injected into the peritoneum of a host capable of accepting said clone to produce antibody containing ascitic fluid; and said ascitic fluid is harvested.
Claim 11
Independent Claim
43 words
A method according to any one of claims 6 or 7, wherein said clones producing said desired immunoglobulin are injected into a host capable of accepting said clone, and including the additional step of extracting said immunoglobulin from the blood of said host.
Claim 12
Independent Claim
16 words
A method according to any one of claims 6 or 7, wherein said immunoglobulin is IgG.
Claim 13
Independent Claim
16 words
A method according to any one of claims 6 or 7, wherein said immunoglobulin is IgM.
Claim 14
Independent Claim
16 words
A method according to any one of claims 6 or 7, wherein said immunoglobulin is IgA.
Claim 15
Independent Claim
16 words
A method according to any one of claims 6 or 7, wherein said immunoglobulin is IgE.
Claim 16
Independent Claim
16 words
A method according to any one of claims 6 or 7, wherein said immunoglobulin is IgD.
Claim 17
Independent Claim
111 words
A method for producing immunoglobulin ("Ig") specific for a predetermined determinant site, said method comprising: co-cultivating in HAT selective conditions HGPRT-deficient EBV-transformed cells as transferring cells and B-lymphocytes activated by an immunogen containing said determinant site as recipient cells for sufficient incubation times and conditions, resulting in the transformation of said recipient cells to immortalized Ig producing cells which are EBNA-positive and which have the normal diploid number of chromosomes, and the mortality of said HGPRT-deficient cells; growing resulting viable cells in HAT medium under limiting dilution conditions and selecting for clones producing immunoglobulin specific for said determinant site; and expanding selected clones and isolating said immunoglobulin secreted by said clones.
Claim 18
Independent Claim
111 words
A method for producing immunoglobulin ("Ig") specific for a predetermined determinant site, said method comprising: co-cultivating in HAT selective conditions HGPRT-deficient EBV-transformed cells as transferring cells and B-lymphocytes activated by an immunogen containing said determinant site as recipient cells for sufficient incubation times and conditions, resulting in the transformation of said recipient cells to immortalized Ig producing cells which are EBNA-positive and which have the normal diploid number of chromosomes, and the mortality of said HGPRT-deficient cells; growing resulting viable cells in HAT medium under limiting dilution conditions and selecting for clones producing immunoglobulin specific for said determinant site; and expanding selected clones and isolating said immunoglobulin secreted by said clones.
Claim 19
Dependent Claim
14 words
A method according to claim 15, wherein said transferring cell is 1A2 (CRL 8119).
Claim 20
Dependent Claim
11 words
A method according to claim 15, wherein said immunoglobulin is IgG.
Claim 21
Dependent Claim
9 words
A human EBV transformed cell designated 1A2 (CRL 8119).
Claim 22
Dependent Claim
9 words
A human EBV transformed cell designated 1A2 (CRL 8119).
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1987-08-07 | 1988-02-06 | 1988-08-07 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1991-08-07 | 1992-02-06 | 1992-08-07 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1995-08-07 | 1996-02-06 | 1996-08-07 | $7700.00 | Window Closed / Lapsed |