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US4808151A
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Simplified method for the preparation of human lymphokine activated killer cells
Assignee: Terumo Corp
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Filing: 1987-04-27
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Granted: 1989-02-28
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Expires: 2007-04-27
Abstract
Abstract A lymphocyte-containing white blood cell fraction obtained by standard leukapheresis, elutriation leukapheresis or standard centrifugation can be used for production of lymphokine activated killer cells by incubation with IL-2. Removal of red blood cells and granulocytes by centrifugation on ficoll is not necessary.
Inventors
- 👤 George F. Dunn, Jr.
- 👤 Joseph D. Irr
- 👤 Lise N. Halpern
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-28 19:51
CPC Classifications
C; C12; C12N; C12N5/00; C12N5/06; C12N5/0602; C12N5/0634; C12N5/0636; A; A61; A61K; A61K40/00; A61K40/10; A61K40/11; A61K40/40; A61K40/41; A61K40/42; C12N5/0081; C12N5/0087; A61K35/00; A61K35/12; A61K2035/124; C12N2500/00; C12N2500/30; C12N2500/32; C12N2501/00; C12N2501/20; C12N2501/23
Claims Analysis
Total Claims: 19
Claim 1
Independent Claim
70 words
In the method of producing lymphokine activated killer cells in vitro which comprises removing red blood cells and plasma from whole blood to produce a lymphocyte-containing white blood cell-rich fraction and incubating the white blood cell-rich fraction in culture medium with interleukin- 2, the improvement which comprises removing red blood cells and plasma and using the white blood cell-rich fraction without an intermediate separation of lymphocytes on a ficoll gradient.
Claim 2
Dependent Claim
70 words
In the method of producing lymphokine activated killer cells in vitro which comprises removing red blood cells and plasma from whole blood to produce a lymphocyte-containing white blood cell-rich fraction and incubating the white blood cell-rich fraction in culture medium with interleukin- 2, the improvement which comprises removing red blood cells and plasma and using the white blood cell-rich fraction without an intermediate separation of lymphocytes on a ficoll gradient.
Claim 3
Dependent Claim
34 words
Method of claim 1 wherein the red blood cells are removed by leukapheresis and the volume percent of red blood cells in the white blood cell-rich fraction is in the range of about 1-20.
Claim 4
Dependent Claim
27 words
Method of claim 2 wherein the red blood cell to white blood cell ratio in the white blood cell-rich fraction is in the range of about 0.2-250.
Claim 5
Dependent Claim
21 words
Method of claim 2 wherein the white blood cell-rich fraction is washed with salt solution prior to incubation to inhibit clotting.
Claim 6
Dependent Claim
34 words
Method of claim 1 wherein the red blood cells are removed by elutriation leukapheresis and the volume percent of red blood cells in the white cell-rich fraction is in the range of about 1-6.
Claim 7
Dependent Claim
27 words
Method of claim 5 wherein the red blood cell to white blood cell ratio in the white blood cell-rich fraction is in the range of about 0.2-50.
Claim 8
Dependent Claim
24 words
Method of claim 6 wherein the white blood cell fraction has a differential of about 80-85% lymphocytes, about 10-20% monocytes, and about 1-5% granulocytes.
Claim 9
Dependent Claim
47 words
Method of claim 7 wherein the volume percent red blood cells in the white blood cell-rich fraction is in the range of about 2-4, and the red blood cell to white blood cell ratio in the white blood cell-rich fraction is in the range of about 0.5-25.
Claim 10
Dependent Claim
23 words
Method of claim 1 wherein monocytes are depleted by treatment with phenyl alanine methyl ester before incubation of the white blood cell-rich fraction.
Claim 11
Dependent Claim
65 words
In the method of generating lymphokine activated killer cells by incubating a lymphocyte-containing white blood cell-rich fraction in culture medium with interleukin-2, the improvement which comprises incubating with interleukin-2 a lymphoctye-containing white blood cell-rich fraction having a red blood cell to white blood cell ratio by number in the range of about 0.2 to 300 and a red blood cell volume percent of about 1-50.
Claim 12
Dependent Claim
65 words
In the method of generating lymphokine activated killer cells by incubating a lymphocyte-containing white blood cell-rich fraction in culture medium with interleukin-2, the improvement which comprises incubating with interleukin-2 a lymphoctye-containing white blood cell-rich fraction having a red blood cell to white blood cell ratio by number in the range of about 0.2 to 300 and a red blood cell volume percent of about 1-50.
Claim 13
Dependent Claim
41 words
Method of claim 10 wherein the red blood cell to white blood cell ratio is in the range of about 0.2-250 and the red blood cell volume percent is in the range of about 1-20 in the white blood cell-rich fraction.
Claim 14
Dependent Claim
41 words
Method of claim 11 wherein the red blood cell to white blood cell ratio is in the range of about 0.2-50 and the red blood cell volume percent is in the range of about 1-6 in the white blood cell-rich fraction.
Claim 15
Dependent Claim
24 words
Method of claim 12 wherein white blood cell-rich fraction has a differential of about 1-5% granulocytes, 0-20% monocytes and greater than about 80% lymphocytes.
Claim 16
Dependent Claim
37 words
Method of claim 13 wherein the red blood cell to white blood cell ratio is in the range of about 0.5-25 and the red blood cell volume content is about 2-4 in the white blood cell-rich fraction.
Claim 17
Dependent Claim
89 words
In the method of treatment of a cancer patient by adoptive immunotherapy which comprises removing pg,43 peripheral blood from the patient, separating a lymphocyte-containing white blood cell-rich fraction from the blood, incubating the lymphocyte-containing white blood cell-rich fraction with interleukin-2 to produce lymphokine-activated killer cells, and reinjecting the activated cells into the patient, the improvement which comprises separating the lymphocyte-containing WBC-rich fraction without use of a ficoll gradient, whereby the volume percent of red blood cells in the white blood cell-rich fraction is in the range of about 1-20.
Claim 18
Dependent Claim
89 words
In the method of treatment of a cancer patient by adoptive immunotherapy which comprises removing pg,43 peripheral blood from the patient, separating a lymphocyte-containing white blood cell-rich fraction from the blood, incubating the lymphocyte-containing white blood cell-rich fraction with interleukin-2 to produce lymphokine-activated killer cells, and reinjecting the activated cells into the patient, the improvement which comprises separating the lymphocyte-containing WBC-rich fraction without use of a ficoll gradient, whereby the volume percent of red blood cells in the white blood cell-rich fraction is in the range of about 1-20.
Claim 19
Dependent Claim
37 words
Method of claim 15 wherein the lymphocyte-containing white blood cell-rich fraction is separated by elutriation leukapheresis whereby the volume percent of red blood cells in the white blood cell fraction is in the range of about 1-6.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1992-02-28 | 1992-08-29 | 1993-02-28 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1996-02-28 | 1996-08-29 | 1997-02-28 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2000-02-28 | 2000-08-29 | 2001-02-28 | $7700.00 | Window Closed / Lapsed |