Recombinant Anti-CD20 antibody [EP459Y] (ab78237)
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
- Suitable for: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-P
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-CD20 antibody [EP459Y]
See all CD20 primary antibodies -
Description
Rabbit monoclonal [EP459Y] to CD20 -
Host species
Rabbit -
Specificity
For optimal performance in IHC, the primary antibody should be incubated overnight at 4?.
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Tested applications
Suitable for: Flow Cyt (Intra), ICC/IF, WB, IP, IHC-Pmore details -
Species reactivity
Reacts with: Human
Predicted to work with: MonkeyDoes not react with: Mouse, Rat -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- IHC-P: Human B cell lymphoma tissue. Human spleen and tonsil tissue. IP: Ramos whole cell lysate. WB: Raji whole cell lysate; Wild-type Raji cell lysate ICC/IF: Ramos cells Flow Cyt (intra): Ramos cells
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General notes
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.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EP459Y -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
- Anti-CD20 antibody [EP459Y] - Low endotoxin, Azide free (ab166865)
- 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)
- Anti-CD20 antibody [EP459Y] - Mouse IgG1 (Chimeric) (ab279298)
- Anti-CD20 antibody [EP459Y] - Mouse IgG2a (Chimeric) (ab279299)
- Anti-CD20 antibody [EP459Y] - Rat IgG2a (Chimeric) (ab279300)
- Alexa Fluor® 568 Anti-CD20 antibody [EP459Y] (ab312637)
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Compatible Secondaries
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Isotype control
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Positive Controls
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab78237 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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Flow Cyt (Intra) |
1/200.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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ICC/IF | (1) |
1/10.
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WB |
1/1000 - 1/10000. Detects a band of approximately 33 kDa (predicted molecular weight: 33 kDa).
|
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IP |
1/20.
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IHC-P | (5) |
1/50 - 1/250. 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?. |
Notes |
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Flow Cyt (Intra)
1/200. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
ICC/IF
1/10. |
WB
1/1000 - 1/10000. Detects a band of approximately 33 kDa (predicted molecular weight: 33 kDa). |
IP
1/20. |
IHC-P
1/50 - 1/250. 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?. |
Target
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Function
This protein may be involved in the regulation of B-cell activation and proliferation. -
Tissue specificity
Expressed on B-cells. -
Involvement in disease
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. -
Sequence similarities
Belongs to the MS4A family. -
Post-translational
modificationsPhosphorylated. Might be functionally regulated by protein kinase(s). -
Cellular localization
Membrane. - Information by UniProt
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Database links
- Entrez Gene: 931 Human
- Omim: 112210 Human
- SwissProt: P11836 Human
- Unigene: 712553 Human
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Alternative names
- APY antibody
- ATOPY antibody
- B lymphocyte antigen CD20 antibody
see all
Images
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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.
Predicted band size: 33 kDa
Observed band size: 33 kDaFalse 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.
Predicted band size: 33 kDa
Observed band size: 33 kDaFalse 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.
Predicted band size: 33 kDa
Observed band size: 33 kDaFalse 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)
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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 Fluorr® 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)
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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 counterstained 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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Formaldehyde-fixed, paraffin-embedded human spleen tissue stained for CD20 using ab78237 at 1/50 dilution in immunohistochemical analysis.
Heat mediated antigen retieval with citrate pH 6.0.
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ab78237 (purified) at 1/20 immunoprecipitating CD20 in 10 μg Ramos (Human Burkitt's lymphoma cell line) 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/Dilution buffer and concentration: 5% NFDM/TBST.
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Unpurified ab78237 showing positive staining in human spleen tissue.
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Unpurified ab78237 showing positive staining in human tonsil tissue.
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Anti-CD20 antibody [EP459Y] (ab78237) at 1/10000 dilution (purified) + Raji (Human Burkitt's lymphoma cell line) whole cell lysate
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 33 kDa
Observed band size: 33 kDaBlocking/Dilution buffer: 5% NFDM/TBST.
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Anti-CD20 antibody [EP459Y] (ab78237) at 1/5000 dilution (unpurified) + Raji (Human Burkitt's lymphoma cell line) cell lysate
Predicted band size: 33 kDa
Observed band size: 33 kDa -
Unpurified ab78237 staining human CD20 in human lymphoma tissue by immunohistochemistry using formalin fixed, paraffin embedded tissue.
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Unpurified ab78237 showing negative staining in human brain tissue.
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Unpurified ab78237 showing negative staining in human heart tissue.
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Unpurified ab78237 showing negative staining in human kidney tissue.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (59)
ab78237 has been referenced in 59 publications.
- Nakamura M et al. Tertiary Lymphoid Structures and Chemokine Landscape in Virus-Positive and Virus-Negative Merkel Cell Carcinoma. Front Oncol 12:811586 (2022). PubMed: 35223493
- Liu X et al. Single-cell transcriptomics links malignant T cells to the tumor immune landscape in cutaneous T cell lymphoma. Nat Commun 13:1158 (2022). PubMed: 35241665
- Song G et al. Single-cell transcriptomic analysis suggests two molecularly subtypes of intrahepatic cholangiocarcinoma. Nat Commun 13:1642 (2022). PubMed: 35347134
- Yu JS et al. The association of immune cell infiltration and prognostic value of tertiary lymphoid structures in gastric cancer. Neoplasma 69:886-898 (2022). PubMed: 35603954
- Liu J et al. Multicenter phase II trial of Camrelizumab combined with Apatinib and Eribulin in heavily pretreated patients with advanced triple-negative breast cancer. Nat Commun 13:3011 (2022). PubMed: 35641481