CA2114311A1 - Monoclonal WB B-Cell Line - Google Patents
Monoclonal WB B-Cell LineInfo
- Publication number
- CA2114311A1 CA2114311A1 CA2114311A CA2114311A CA2114311A1 CA 2114311 A1 CA2114311 A1 CA 2114311A1 CA 2114311 A CA2114311 A CA 2114311A CA 2114311 A CA2114311 A CA 2114311A CA 2114311 A1 CA2114311 A1 CA 2114311A1
- Authority
- CA
- Canada
- Prior art keywords
- cells
- protein
- cell
- cell line
- magnification
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 210000003719 b-lymphocyte Anatomy 0.000 title abstract 3
- 210000004027 cell Anatomy 0.000 abstract 15
- 102000004169 proteins and genes Human genes 0.000 abstract 6
- 108090000623 proteins and genes Proteins 0.000 abstract 6
- 238000003917 TEM image Methods 0.000 abstract 4
- 210000003935 rough endoplasmic reticulum Anatomy 0.000 abstract 4
- 210000003000 inclusion body Anatomy 0.000 abstract 3
- 210000003470 mitochondria Anatomy 0.000 abstract 3
- 102100031585 ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase 1 Human genes 0.000 abstract 2
- 101000777636 Homo sapiens ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase 1 Proteins 0.000 abstract 2
- 101001057504 Homo sapiens Interferon-stimulated gene 20 kDa protein Proteins 0.000 abstract 2
- 101001055144 Homo sapiens Interleukin-2 receptor subunit alpha Proteins 0.000 abstract 2
- 101001018097 Homo sapiens L-selectin Proteins 0.000 abstract 2
- 101000878605 Homo sapiens Low affinity immunoglobulin epsilon Fc receptor Proteins 0.000 abstract 2
- 101000835093 Homo sapiens Transferrin receptor protein 1 Proteins 0.000 abstract 2
- 102100026878 Interleukin-2 receptor subunit alpha Human genes 0.000 abstract 2
- 102100033467 L-selectin Human genes 0.000 abstract 2
- 102100038007 Low affinity immunoglobulin epsilon Fc receptor Human genes 0.000 abstract 2
- 102100026144 Transferrin receptor protein 1 Human genes 0.000 abstract 2
- 150000001720 carbohydrates Chemical class 0.000 abstract 2
- 239000003636 conditioned culture medium Substances 0.000 abstract 2
- 238000001493 electron microscopy Methods 0.000 abstract 2
- 230000001788 irregular Effects 0.000 abstract 2
- 102100024222 B-lymphocyte antigen CD19 Human genes 0.000 abstract 1
- 102100022005 B-lymphocyte antigen CD20 Human genes 0.000 abstract 1
- 101000980825 Homo sapiens B-lymphocyte antigen CD19 Proteins 0.000 abstract 1
- 101000897405 Homo sapiens B-lymphocyte antigen CD20 Proteins 0.000 abstract 1
- 206010028980 Neoplasm Diseases 0.000 abstract 1
- 239000012980 RPMI-1640 medium Substances 0.000 abstract 1
- 102000040739 Secretory proteins Human genes 0.000 abstract 1
- 108091058545 Secretory proteins Proteins 0.000 abstract 1
- 230000004913 activation Effects 0.000 abstract 1
- 239000000427 antigen Substances 0.000 abstract 1
- 102000036639 antigens Human genes 0.000 abstract 1
- 108091007433 antigens Proteins 0.000 abstract 1
- 230000006907 apoptotic process Effects 0.000 abstract 1
- 201000011510 cancer Diseases 0.000 abstract 1
- 230000001086 cytosolic effect Effects 0.000 abstract 1
- 239000008187 granular material Substances 0.000 abstract 1
- 238000001727 in vivo Methods 0.000 abstract 1
- 238000011534 incubation Methods 0.000 abstract 1
- 238000004519 manufacturing process Methods 0.000 abstract 1
- 239000002609 medium Substances 0.000 abstract 1
- 230000001338 necrotic effect Effects 0.000 abstract 1
- 210000005259 peripheral blood Anatomy 0.000 abstract 1
- 239000011886 peripheral blood Substances 0.000 abstract 1
- 238000002135 phase contrast microscopy Methods 0.000 abstract 1
- 230000035755 proliferation Effects 0.000 abstract 1
- 238000000746 purification Methods 0.000 abstract 1
- 230000010076 replication Effects 0.000 abstract 1
- 238000004114 suspension culture Methods 0.000 abstract 1
- 210000003934 vacuole Anatomy 0.000 abstract 1
- 230000003442 weekly effect Effects 0.000 abstract 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0634—Cells from the blood or the immune system
- C12N5/0635—B lymphocytes
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/90—Serum-free medium, which may still contain naturally-sourced components
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/90—Serum-free medium, which may still contain naturally-sourced components
- C12N2500/95—Protein-free medium and culture conditions
Abstract
A human B-cell line (designated WB) was selectively cloned from peripheral blood of a healthy 48 year-old Chinese male by use of a serum-free defined medium. The in vivo pre-activated B-cells required an 8-week "incubation period" before large clusters of cells with apical cytoplasmic projections directed radially outwards became apparent, and their gradual proliferation was observed. Within a week, sufficient cell clusters predominate to allow a 1:2 split twice weekly. Phase-contrast microscopy showed individual cells as 10u to 20u fusiforms with abundant phase-dense granules. By electron microscopy, each cell revealed numerous inclusion bodies, mitochondria, stratified rough endoplasmic reticulum (RER), carbohydrate inclusions, and a heterochromatic thick-rimmed, irregular large nucleus containing a prominent nucleolus. These EBV-negative WB cells expressed the CD19 and CD20 pan-B antigens, IgG, and activation markers DR, DQ, Leu 8, CD23, CD25, CD38, and CD71.
After 6 months of subculture, the cell line was adapted to culture in a protein-free RPMI
1640 medium. Interestingly, the cells could sustain autonomous growth and replication in semi-suspension culture in the absence of any exogenous protein. Under electron microscopy, these protein-independent cells exhibited ringed nuclei, frequent binucleism, dense elongated mitochondria, and carbohydrate inclusions. In addition, there was abrogation of DR, DQ, Leu 8, CD23, CD25, CD38, and CD71. The conditioned medium (CM) from either the serum-free or the protein-free WB culture contained two novel proteins, namely 21 and 53 kilodaltons (Kd) moeities with respective necrotic and growth bioactivity. Hence, availability of this cell line would allow for easy purification of these important secretory proteins, and for the production of human monoclonal antibodies. Also, further study of the cell line may provide the answer as to how cancer cells could gain autonomy in a milieu of apoptosis.
DESCRIPTION OF FIGURES
Figure 1.
Transmission-electron micrograph of a representative WB cell showing inclusion bodies, stratified rough endoplasmic reticulum (RER), and a heterochromatic thicked-rimmed, irregular large nucleus with a prominent nucleolus. Magnification: 26,000X.
Figure 2.
Transmission-electron micrograph of a ringed nucleus surrounding a central vacuole displayed by an occasional protein-independent WB cell. Magnification: 26,000X.
Figure 3.
Transmission-electron micrograph of a representative binucleism shown by several protein-independent WB cells. Magnification: 26,000X.
Figure 4.
Transmission-electron micrograph of dense elongated mitochondria and inclusion bodies found in the protein-independent WB cell. Magnification: 26,000X.
After 6 months of subculture, the cell line was adapted to culture in a protein-free RPMI
1640 medium. Interestingly, the cells could sustain autonomous growth and replication in semi-suspension culture in the absence of any exogenous protein. Under electron microscopy, these protein-independent cells exhibited ringed nuclei, frequent binucleism, dense elongated mitochondria, and carbohydrate inclusions. In addition, there was abrogation of DR, DQ, Leu 8, CD23, CD25, CD38, and CD71. The conditioned medium (CM) from either the serum-free or the protein-free WB culture contained two novel proteins, namely 21 and 53 kilodaltons (Kd) moeities with respective necrotic and growth bioactivity. Hence, availability of this cell line would allow for easy purification of these important secretory proteins, and for the production of human monoclonal antibodies. Also, further study of the cell line may provide the answer as to how cancer cells could gain autonomy in a milieu of apoptosis.
DESCRIPTION OF FIGURES
Figure 1.
Transmission-electron micrograph of a representative WB cell showing inclusion bodies, stratified rough endoplasmic reticulum (RER), and a heterochromatic thicked-rimmed, irregular large nucleus with a prominent nucleolus. Magnification: 26,000X.
Figure 2.
Transmission-electron micrograph of a ringed nucleus surrounding a central vacuole displayed by an occasional protein-independent WB cell. Magnification: 26,000X.
Figure 3.
Transmission-electron micrograph of a representative binucleism shown by several protein-independent WB cells. Magnification: 26,000X.
Figure 4.
Transmission-electron micrograph of dense elongated mitochondria and inclusion bodies found in the protein-independent WB cell. Magnification: 26,000X.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA002114311A CA2114311C (en) | 1991-11-25 | 1991-11-25 | Monoclonal wb b-cell line |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA002114311A CA2114311C (en) | 1991-11-25 | 1991-11-25 | Monoclonal wb b-cell line |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2114311A1 true CA2114311A1 (en) | 1993-05-26 |
| CA2114311C CA2114311C (en) | 1996-02-27 |
Family
ID=4152799
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002114311A Expired - Fee Related CA2114311C (en) | 1991-11-25 | 1991-11-25 | Monoclonal wb b-cell line |
Country Status (1)
| Country | Link |
|---|---|
| CA (1) | CA2114311C (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0792278A1 (en) * | 1994-11-07 | 1997-09-03 | Human Genome Sciences, Inc. | Tumor necrosis factor-gamma |
| US6599719B2 (en) | 1994-11-07 | 2003-07-29 | Human Genome Sciences, Inc. | Nucleic acid molecules encoding tumor necrosis factor-gamma-alpha |
| US6824767B2 (en) | 1994-11-07 | 2004-11-30 | Human Genome Sciences, Inc. | Tumor necrosis factor-gamma |
-
1991
- 1991-11-25 CA CA002114311A patent/CA2114311C/en not_active Expired - Fee Related
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0792278A1 (en) * | 1994-11-07 | 1997-09-03 | Human Genome Sciences, Inc. | Tumor necrosis factor-gamma |
| EP0792278A4 (en) * | 1994-11-07 | 1999-09-22 | Human Genome Sciences Inc | Tumor necrosis factor-gamma |
| US6599719B2 (en) | 1994-11-07 | 2003-07-29 | Human Genome Sciences, Inc. | Nucleic acid molecules encoding tumor necrosis factor-gamma-alpha |
| US6824767B2 (en) | 1994-11-07 | 2004-11-30 | Human Genome Sciences, Inc. | Tumor necrosis factor-gamma |
Also Published As
| Publication number | Publication date |
|---|---|
| CA2114311C (en) | 1996-02-27 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| EEER | Examination request | ||
| MKLA | Lapsed |