重组Anti-CD20抗体[EP459Y] - Low endotoxin,Azide free (ab166865)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP459Y] to CD20 - Low endotoxin, Azide free
- Suitable for: Flow Cyt (Intra), WB, ICC/IF, IP, IHC-P
- Knockout validated
- Reacts with: Human
概述
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产品名称
Anti-CD20抗体[EP459Y] - Low endotoxin,Azide free
参阅全部 CD20 一抗 -
描述
兔单克隆抗体[EP459Y] to CD20 - Low endotoxin,Azide free -
宿主
Rabbit -
特异性
For optimal performance in IHC, the primary antibody should be incubated overnight at 4?.
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经测试应用
适用于: Flow Cyt (Intra), WB, ICC/IF, IP, IHC-Pmore details -
种属反应性
与反应: Human
预测可用于: Monkey不与反应: Mouse, Rat -
免疫原
Synthetic peptide within Human CD20 aa 250-350 (C terminal) conjugated to keyhole limpet haemocyanin. The exact sequence is proprietary.
Database link: P11836 -
阳性对照
- WB: Wild-type Raji cell lysate Flow Cyt (intra): Ramos cells
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常规说明
ab166865 is the carrier-free version of ab78237.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Our Low endotoxin, azide-free formats have low endotoxin level (≤ 1 EU/ml, determined by the LAL assay) and are free from azide, to achieve consistent experimental results in functional assays.
性能
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形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
存储溶液
Constituent: 100% PBS -
无载体
是 -
Concentration information loading...
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纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EP459Y -
同种型
IgG -
研究领域
相关产品
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Alternative Versions
- Alexa Fluor® 488 Anti-CD20 antibody [EP459Y] (ab198941)
- Alexa Fluor® 647 Anti-CD20 antibody [EP459Y] (ab198943)
- Anti-CD20 antibody [EP459Y] - BSA and Azide free (ab214282)
- PE Anti-CD20 antibody [EP459Y] (ab303031)
- APC Anti-CD20 antibody [EP459Y] (ab303032)
- HRP Anti-CD20 antibody [EP459Y] (ab303033)
- Alexa Fluor® 594 Anti-CD20 antibody [EP459Y] (ab310623)
- Alexa Fluor® 555 Anti-CD20 antibody [EP459Y] (ab312151)
- Alexa Fluor® 568 Anti-CD20 antibody [EP459Y] (ab312637)
- Anti-CD20 antibody [EP459Y] (ab78237)
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Compatible Secondaries
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Conjugation kits
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Isotype control
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab166865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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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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WB |
1/1000.
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ICC/IF |
Use at an assay dependent concentration.
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IP |
Use at an assay dependent concentration.
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
For optimal performance in IHC, the primary antibody should be incubated overnight at 4?. |
说明 |
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Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
1/1000. |
ICC/IF
Use at an assay dependent concentration. |
IP
Use at an assay dependent concentration. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. For optimal performance in IHC, the primary antibody should be incubated overnight at 4?. |
靶标
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功能
This protein may be involved in the regulation of B-cell activation and proliferation. -
组织特异性
Expressed on B-cells. -
疾病相关
Defects in MS4A1 are the cause of immunodeficiency common variable type 5 (CVID5) [MIM:613495]; also called antibody deficiency due to CD20 defect. CVID5 is a primary immunodeficiency characterized by antibody deficiency, hypogammaglobulinemia, recurrent bacterial infections and an inability to mount an antibody response to antigen. The defect results from a failure of B-cell differentiation and impaired secretion of immunoglobulins; the numbers of circulating B cells is usually in the normal range, but can be low. -
序列相似性
Belongs to the MS4A family. -
翻译后修饰
Phosphorylated. Might be functionally regulated by protein kinase(s). -
细胞定位
Membrane. - Information by UniProt
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数据库链接
- Entrez Gene: 931 Human
- Omim: 112210 Human
- SwissProt: P11836 Human
- Unigene: 712553 Human
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别名
- APY antibody
- ATOPY antibody
- B lymphocyte antigen CD20 antibody
see all
图片
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab78237).
Flow cytometry overlay histogram showing left Ramos positive cells and right negative HEK293 stained with ab78237 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab78237) (1x 106 in 100μl at 0.2μg/ml (1/2500)) for 30min at 22°C.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.
This antibody gave a positive signal in Ramos Fixed with 80% methanol (5 min) / permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
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All lanes : Anti-CD20 antibody [EP459Y] - Rat IgG2a (Chimeric) (ab279300) at 1/1000 dilution
Lane 1 : Wild-type Raji cell lysate
Lane 2 : MS4A1 knockout Raji cell lysate
Lane 3 : A549 cell lysate
Lane 4 : HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 33 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-CD20 antibody [EP459Y] - Rat IgG2a staining at 1/1000 dilution, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab279300 was shown to bind specifically to CD20. A band was observed at 33 kDa in wild-type Raji cell lysates with no signal observed at this size in MS4A1 knockout cell line ab273871 (knockout cell lysate ab263259). To generate this image, wild-type and MS4A1 knockout Raji cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rat IgG H&L (IRDye® 800CW) preabsorbed (ab253031) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.
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All lanes : Anti-CD20 antibody [EP459Y] - Mouse IgG2a (Chimeric) (ab279299) at 1/1000 dilution
Lane 1 : Wild-type Raji cell lysate
Lane 2 : MS4A1 knockout Raji cell lysate
Lane 3 : A549 cell lysate
Lane 4 : HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 33 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-CD20 antibody [EP459Y] - Mouse IgG2a staining at 1/1000 dilution, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab279299 was shown to bind specifically to CD20. A band was observed at 33 kDa in wild-type Raji cell lysates with no signal observed at this size in MS4A1 knockout cell line ab273871 (knockout cell lysate ab263259). To generate this image, wild-type and MS4A1 knockout Raji cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.
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All lanes : Anti-CD20 antibody [EP459Y] - Mouse IgG1 (Chimeric) (ab279298) at 1/1000 dilution
Lane 1 : Wild-type Raji cell lysate
Lane 2 : MS4A1 knockout Raji cell lysate
Lane 3 : A549 cell lysate
Lane 4 : HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 33 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-CD20 antibody [EP459Y] - Mouse IgG1 staining at 1/1000 dilution, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab279298 was shown to bind specifically to CD20. A band was observed at 33 kDa in wild-type Raji cell lysates with no signal observed at this size in MS4A1 knockout cell line ab273871 (knockout cell lysate ab263259). To generate this image, wild-type and MS4A1 knockout Raji cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) at 1/20000 dilution.
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Immunocytochemistry/ Immunofluorescence analysis of Ramos (human Burkitt's lymphoma B lymphocyte) labeling CD20 with purified ab78237 at 1/10 dilution. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/1000 was used as the secondary antibody. Cells were counterstained with ab195889, Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200. PBS instead of the primary antibody was used as negative control. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Nuclei were stained with DAPI (blue).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab78237).
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This IHC data was generated using the same anti-CD20 antibody clone, EP459Y, in a different buffer formulation (cat# ab78237).
Immunohistochemical staining of paraffin embedded human B cell lymphoma with purified ab78237 at a working dilution of 1/50. The secondary antibody used is ab97051, a goat anti-rabbit IgG (H&L) at a dilution of 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.
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Intracellular Flow Cytometry analysis ofRamos (Human Burkitt's lymphoma B lymphocyte) labeling CD20 with purified ab78237 at 1/200 dilution (Red). Goat anti rabbit IgG (Alexa Fluor®488, ab150081) at 1/2000 dilution was used as secondary antibody. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol.
Isotype control: Rabbit monoclonal IgG (ab172730) (Black)
Unlabelled cells: (Blue)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab78237).
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ab78237 (purified) at 1/20 immunoprecipitating CD20 in 10 µg Ramos cell lysate (Lanes 1 and 2, observed at 33 kDa). Lane 3 - Rabbit monoclonal IgG (ab172730). For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution. Blocking buffer and concentration: 5% NFDM/TBST Dilution buffer and concentration: 5% NFDM/TBST
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab78237).
实验方案
数据表及文件
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Datasheet download
文献 (2)
ab166865 被引用在 2 文献中.
- Cheung AM et al. Quantitative single-cell analysis of immunofluorescence protein multiplex images illustrates biomarker spatial heterogeneity within breast cancer subtypes. Breast Cancer Res 23:114 (2021). PubMed: 34922607
- Xu-Monette ZY et al. Immune Profiling and Quantitative Analysis Decipher the Clinical Role of Immune-Checkpoint Expression in the Tumor Immune Microenvironment of DLBCL. Cancer Immunol Res 7:644-657 (2019). PubMed: 30745366