人PDGFRB knockout SH-SY5Y cell line (ab273749)
概述
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产品名称
人PDGFRB knockout SH-SY5Y cell line
参阅全部 PDGFR beta 细胞裂解液 -
Parental Cell Line
SHSY-5Y -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 5 bp deletion in exon 3 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
经测试应用
适用于: WBmore details -
Biosafety level
1 -
常规说明
Recommended control: Human wild-type SHSY-5Y cell line (ab275475). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: 1:1 mixture of EMEM and F-12K + 10% FBS
Initial handling guidelines: Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method. Based on cell count, seed cells in an appropriate cell culture flask at a density of 1-2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules.
4. Incubate the culture at 37°C incubator with 5% CO2. Cultures should be monitored daily.Subculture guidelines:
- All seeding densities should be based on cell counts gained by established methods.
- These cells grow as a mixture of floating and adherent cells.
- Remove media containing floating cells and recover cells by centrifugation, detach cells using standard methods, combine with floating cells and transfer to a new culture flask.
- A guide seeding density of 1-2x104 cells/cm2 is recommended.
- Cells should be seeded at a density conducive to cell–cell communication to proliferate. If cells are seeded too sparsely, growth rate is reduced and cell death is high.
This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
We will provide viable cells that proliferate on revival.
性能
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Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Bone marrow -
Cell type
neuroblastoma -
Disease
Neuroblastoma -
Gender
Female -
Mycoplasma free
Yes -
存放说明
Shipped on Dry Ice. Store in liquid nitrogen. -
存储溶液
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
研究领域
靶标
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功能
Receptor that binds specifically to PDGFB and PDGFD and has a tyrosine-protein kinase activity. Phosphorylates Tyr residues at the C-terminus of PTPN11 creating a binding site for the SH2 domain of GRB2. -
疾病相关
Note=A chromosomal aberration involving PDGFRB is found in a form of chronic myelomonocytic leukemia (CMML). Translocation t(5;12)(q33;p13) with EVT6/TEL. It is characterized by abnormal clonal myeloid proliferation and by progression to acute myelogenous leukemia (AML).
Note=A chromosomal aberration involving PDGFRB may be a cause of acute myelogenous leukemia. Translocation t(5;14)(q33;q32) with TRIP11. The fusion protein may be involved in clonal evolution of leukemia and eosinophilia.
Note=A chromosomal aberration involving PDGFRB may be a cause of juvenile myelomonocytic leukemia. Translocation t(5;17)(q33;p11.2) with SPECC1.
Defects in PDGFRB are a cause of myeloproliferative disorder chronic with eosinophilia (MPE) [MIM:131440]. A hematologic disorder characterized by malignant eosinophils proliferation. Note=A chromosomal aberration involving PDGFRB is found in many instances of myeloproliferative disorder chronic with eosinophilia. Translocation t(5;12) with ETV6 on chromosome 12 creating an PDGFRB-ETV6 fusion protein.
Note=A chromosomal aberration involving PDGFRB may be the cause of a myeloproliferative disorder (MBD) associated with eosinophilia. Translocation t(1;5)(q23;q33) that forms a PDE4DIP-PDGFRB fusion protein. -
序列相似性
Belongs to the protein kinase superfamily. Tyr protein kinase family. CSF-1/PDGF receptor subfamily.
Contains 5 Ig-like C2-type (immunoglobulin-like) domains.
Contains 1 protein kinase domain. -
翻译后修饰
Autophosphorylated. Dephosphorylated by PTPRJ at Tyr-751, Tyr-857, Tyr-1009 and Tyr-1021. -
细胞定位
Membrane. - Information by UniProt
相关产品
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KO cell lysates
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Related Products
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab273749于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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WB |
Use at an assay dependent concentration.
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说明 |
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WB
Use at an assay dependent concentration. |
图片
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All lanes : Anti-PDGFR beta antibody [42G12] (ab69506) at 1/1000 dilution
Lane 1 : Wild-type SH-SY5Y cell lysate
Lane 2 : PDGFRB knockout SH-SY5Y cell lysate
Lane 3 : Human Skeletal Muscle tissue lysate
Lane 4 : HeLa cell lysate
Lysates/proteins at 30 µg per lane.
Performed under reducing conditions.
Observed band size: 170 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab69506 observed at 170 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55 kDa.
ab69506 was shown to react with PDGFR beta in wild-type SH-SY5Y cells in Western blot with loss of signal observed in PDGFRB knockout cell line ab273749 (knockout cell lysate ab275523). Wild-type SH-SY5Y and PDGFRB knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab69506 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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All lanes : Anti-PDGFR alpha + PDGFR beta antibody [Y92] - C-terminal (ab32570) at 1/5000 dilution
Lane 1 : Wild-type SH-SY5Y cell lysate
Lane 2 : PDGFR beta knockout SH-SY5Y cell lysate
Lane 3 : Human Skeletal Muscle tissue lysate
Lane 4 : HeLa cell lysate
Lysates/proteins at 30 µg per lane.
Performed under reducing conditions.
Observed band size: 170 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab32570 observed at 170 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab32570 was shown to react with PDGFRB in wild-type SH-SY5Y cells in Western blot with loss of signal observed in PDGFRB knockout cell line ab273749 (knockout cell lysate ab275523). Wild-type SH-SY5Y and PDGFRB knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab32570 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 5000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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Allele-1: 5 bp deletion in exon 3
实验方案
数据表及文件
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SDS download
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Datasheet download
文献 (0)
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