CD20抗体[SP32]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD20 antibody [SP32] (AB64088)

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling CD20 with ab64088 at 1/100 dilution.

Positive staining on human tonsil.

The section was incubated with ab64088 for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Counterstained with Hematoxylin.

Heat mediated antigen retrieval was performed with Sodium-Citrate buffer (pH 6.0, Epitope Retrieval Solution1) for 10 mins.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-CD20 antibody [SP32] (AB64088)

Flow cytometry analysis of Ramos (human Burkitt's lymphoma) labeling CD20 with purified ab64088 at 1/40 dilution (7.78 μg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotype control -Rabbit monoclonal IgG (ab172730) (Black). Unlableled control -Unlabelled cells (blue).

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human colon tissue staining CD3 with ab21703 at a 1/150 (0.06 ug/ml) dilution, ab318205 anti-TCR beta used at 1/500 (1.036 ug/ml) dilution and ab64088 anti-CD20 used at a 1/50 (0.2 ug/ml) dilution.

Panel A : merged staining of anti-CD3 (magenta; Opal™690), anti-TCR beta (green; Opal™520) and anti-CD20 (yellow; Opal™570) on human colon.
Panel B : anti-CD3 staining T lymphocytes in human colon.
Panel C : anti-TCR beta staining T lymphocytes in human colon.
Panel D : anti-CD20 staining B lymphocytes in human colon.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab21703, ab318205 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human tonsil tissue staining CD3 with ab21703 at a 1/150 (0.06 ug/ml) dilution, ab318205 anti-TCR beta used at 1/500 (1.036 ug/ml) dilution and ab64088 anti-CD20 used at a 1/50 (0.2 ug/ml) dilution.

Panel A : merged staining of anti-CD3 (magenta; Opal™690), anti-TCR beta (green; Opal™520) and anti-CD20 (yellow; Opal™570) on human tonsil.
Panel B : anti-CD3 staining T lymphocytes in human tonsil.
Panel C : anti-TCR beta staining T lymphocytes in human tonsil.
Panel D : anti-CD20 staining B lymphocytes in human tonsil.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab21703, ab318205 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human spleen tissue staining CD3 with ab21703 at a 1/150 (0.06 ug/ml) dilution, ab318205 anti-TCR beta used at 1/500 (1.036 ug/ml) dilution and ab64088 anti-CD20 used at a 1/50 (0.2 ug/ml) dilution.

Panel A : merged staining of anti-CD3 (magenta; Opal™690), anti-TCR beta (green; Opal™520) and anti-CD20 (yellow; Opal™570) on human spleen.
Panel B : anti-CD3 staining T lymphocytes in human spleen.
Panel C : anti-TCR beta staining T lymphocytes in human spleen.
Panel D : anti-CD20 staining B lymphocytes in human spleen.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab21703, ab318205 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-CD20 antibody [SP32] (AB64088)

Flow cytometric analysis of rabbit anti-CD20 (SP32) antibody ab64088(1/100) in Jurkat cells (green) compared to negative control of rabbit IgG (blue).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-CD20 antibody [SP32] (AB64088)

Immunocytochemistry/ Immunofluorescence analysis of Ramos (human Burkitt's lymphoma B lymphocyte) cells labeling CD20 with purified ab64088 at 1/100 (3.1 μg/ml). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 μg/ml). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/ml) dilution. DAPI was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Collaborator

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD20 antibody [SP32] (AB64088)

The BOND™ Polymer Refine Detection System on the BOND RX Automated Stainer (DS9800, Leica Biosystems, Wetzlar, Germany) was used for immunohistochemical labelling of formalin-fixed, paraffin-embedded (FFPE) tissues. The protocol includes a peroxide block, post-primary reagent, polymer detection, DAB chromogen, and haematoxylin counterstain, all automated to minimize variability. Antigen retrieval was performed by heat-induced epitope retrieval (HIER) for 20 minutes in Tris-EDTA buffer (BOND Epitope Retrieval Solution 2, AR9640, Leica Biosystems, Wetzlar, Germany). For mouse and rabbit antibodies, the sequence comprised peroxide block (10 min), primary antibody incubation (30 min), post-primary (10 min), polymer (10 min), DAB (10 min), and haematoxylin (8 min). For rat antibodies, an anti-rat step (1 : 300 dilution, Vector Laboratories, California, USA; 20 min) was included prior to polymer application (15 min).

For mouse tissues, antigen retrieval was performed using high pH buffers (Dako, Agilent), followed by blocking of endogenous peroxidase activity with 3% hydrogen peroxide. Slides were then incubated with the primary antibody. Antigen/antibody complexes detection was carried out using a horseradish peroxidase-conjugated visualization system (Novolink Polymer, Leica). Staining was developed with 3,3′-diaminobenzidine (EnVision FLEX DAB Enhancer, Dako, Agilent), and nuclei were counterstained with hematoxylin (FLEX Hematoxylin, SM806, Dako, Agilent). Images were acquired using an Axiocam CCD camera (Zeiss) and processed with ZEN 2.1 software and Photoshop 9.0.

• IP

Lab

Immunoprecipitation - Anti-CD20 antibody [SP32] (AB64088)

ab64088 at 1/30 dilution immunoprecipitating CD20 in Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate. Western blot was performed from the immunoprecipitate using ab64088 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution. Lane 1 : Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate 10 µg Lane 2 : ab64088 IP in Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab64088 in Raji whole cell lysate Blocking/Dilution buffer : 5% NFDM/TBST. Exposure time : 8 seconds.

Lane 2:

Immunoprecipitation - Anti-CD20 antibody [SP32] (ab64088) at 1/30 dilution

Lane 2:

Immunoprecipitation - Anti-CD20 antibody [SP32] - BSA and Azide free (<a href='/products/primary-antibodies/cd20-antibody-sp32-bsa-and-azide-free-ab236434'>ab236434</a>)

All lanes:

HeLa (human cervix adenocarcinoma epithelial cell) treated with 500 µM CoCl2 for 24 hours whole cell lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 25 kDa,33 kDa

false

Exposure time: 8s

• mIHC

Lab

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Immunohistochemistry analysis of (Formalin/PFA-fixed paraffin-embedded sections) Rat lymph node tissue labelling MAdCAM1 with ab309487 at 1 : 100 dilution (B), CD3 epsilon with ab237721at 1 : 2000 dilution (C) and CD20 with ab64088 at 1 : 100 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-MAdCAM (green; Opal™520), anti-CD3 (gray; Opal™690) and anti-CD20 (magenta; Opal™570) on rat lymph node.

Panel B : anti-MAdCAM staining high endothelial venules in rat lymph node.

Panel C : ant-CD3 staining B lymphocytes in rat lymph node.

Panel D : ant-CD20 staining B lymphocytes in rat lymph node.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab309487, ab237721 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Immunohistochemistry analysis of (Formalin/PFA-fixed paraffin-embedded sections) Mouse lymph node tissue labelling MAdCAM1 with ab309487 at 1 : 100 dilution (B), CD3 epsilon with ab237721at 1 : 2000 dilution (C) and CD20 with ab64088 at 1 : 100 dilution (D). Opal Polymer HRP Ms + Rb was used as a secondary antibody, and DAPI was used for a nuclear counter stain. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

Panel A : merged staining of anti-MAdCAM (green; Opal™520), anti-CD3 (gray; Opal™690) and anti-CD20 (magenta; Opal™570) on mouse lymph node.

Panel B : anti-MAdCAM staining high endothelial venules in mouse lymph node.

Panel C : ant-CD3 staining B lymphocytes in mouse lymph node.

Panel D : ant-CD20 staining B lymphocytes in mouse lymph node.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab309487, ab237721 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-CD20 antibody [SP32] (AB64088)

Flow cytometry analysis of WEHI-231 (mouse lymphoblast B cell lymphoma) labeling CD20 with purified ab64088 at 1/40 dilution (7.78 μg/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as a secondary antibody. Isotype control -Rabbit monoclonal IgG (ab172730) (Black). Unlableled control -Unlabelled cells (blue).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD20 antibody [SP32] (AB64088)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling CD20 with ab64088 at 1/100 dilution.

Positive staining on mouse spleen.

The section was incubated with ab64088 for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Counterstained with Hematoxylin.

Heat mediated antigen retrieval was performed with Sodium-Citrate buffer (pH 6.0, Epitope Retrieval Solution1) for 10 mins.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded rat liver tissue staining MARCO with ab323717 at a 1 : 200 (2.49 ug/ml) dilution, ab64088 anti-CD20 used at 1 : 100 (2.91 ug/ml) dilution and ab237721 anti-CD3 used at a 1 : 2000 (0.26 ug/ml) dilution.

Panel A : merged staining of anti-MARCO (green; Opal™520), anti-CD20 (gray; Opal™690) and anti-CD3 (magenta; Opal™570) on rat liver.
Panel B : anti-MARCO staining macrophages in rat liver.
Panel C : ant-CD20 staining B lymphocytes in rat liver.
Panel D : ant-CD3 staining T lymphocytes in rat liver.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab323717, ab64088 and ab237721 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

This image was generated using ab236434, the same clone, but with a different buffer formulation. Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Rat spleen tissue labeling Sialoadhesin/CD169, CD3 epsilon and CD20 with ab312840 at 1/100 dilution, ab16669 at 1 : 150 dilution and ab236434 at 1 : 5000 dilution. Panel A : merged staining of anti-Sialoadhesin/CD169 (red; Opal™690), anti-CD3 epsilon (green; Opal™520) and anti-CD20 (gray; Opal™570) on rat spleen. Panel B : anti-Sialoadhesin/CD169 stained on macrophages. Panel C : anti-CD3 epsilon stained on T cells. Panel D : anti-CD20 stained on B cells. The section was incubated in three rounds of staining : in the order of ab312840, ab16669, and ab236434 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

This image was generated using ab236434, the same clone, but with a different buffer formulation. Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Mouse spleen tissue labeling Sialoadhesin/CD169, CD3 epsilon and CD20 with ab312840 at 1/100 dilution, ab16669 at 1 : 150 dilution and ab236434 at 1 : 5000 dilution. Panel A : merged staining of anti-Sialoadhesin/CD169 (red; Opal™690), anti-CD3 epsilon (green; Opal™520) and anti-CD20 (gray; Opal™570) on mouse spleen. Panel B : anti-Sialoadhesin/CD169 stained on macrophages. Panel C : anti-CD3 epsilon stained on T cells. Panel D : anti-CD20 stained on B cells. The section was incubated in three rounds of staining : in the order of ab312840, ab16669, and ab236434 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded rat spleen tissue staining CD3 with ab21703 at a 1/150 (0.06 ug/ml) dilution, ab318205 anti-TCR beta used at 1/500 (1.036 ug/ml) dilution and ab64088 anti-CD20 used at a 1/50 (0.2 ug/ml) dilution.

Panel A : merged staining of anti-CD3 (magenta; Opal™690), anti-TCR beta (green; Opal™520) and anti-CD20 (yellow; Opal™570) on rat spleen.
Panel B : anti-CD3 staining T lymphocytes in rat spleen.
Panel C : anti-TCR beta staining T lymphocytes in rat spleen.
Panel D : anti-CD20 staining B lymphocytes in rat spleen.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab21703, ab318205 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• mIHC

Supplier Data

Multiplex immunohistochemistry - Anti-CD20 antibody [SP32] (AB64088)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded mouse spleen tissue staining CD3 with ab21703 at a 1/150 (0.06 ug/ml) dilution, ab318205 anti-TCR beta used at 1/500 (1.036 ug/ml) dilution and ab64088 anti-CD20 used at a 1/50 (0.2 ug/ml) dilution.

Panel A : merged staining of anti-CD3 (magenta; Opal™690), anti-TCR beta (green; Opal™520) and anti-CD20 (yellow; Opal™570) on mouse spleen.
Panel B : anti-CD3 staining T lymphocytes in mouse spleen.
Panel C : anti-TCR beta staining T lymphocytes in mouse spleen.
Panel D : anti-CD20 staining B lymphocytes in mouse spleen.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab21703, ab318205 and ab64088 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Nuclear counter stain with DAPI.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD20 antibody [SP32] (AB64088)

Immunohistochemical analysis of paraffin-embedded Rat spleen tissue labeling CD20 with ab64088 at 1/100 dilution.

Positive staining on rat spleen.

The section was incubated with ab64088 for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Counterstained with Hematoxylin.

Heat mediated antigen retrieval was performed with Sodium-Citrate buffer (pH 6.0, Epitope Retrieval Solution1) for 10 mins.

• WB

Unknown

Western blot - Anti-CD20 antibody [SP32] (AB64088)

All lanes:

Western blot - Anti-CD20 antibody [SP32] (ab64088) at 1/100 dilution

All lanes:

lysate prepared from Ramos cells

Predicted band size: 33 kDa

Observed band size: 33 kDa

false

• WB

Lab

Western blot - Anti-CD20 antibody [SP32] (AB64088)

Western blot : Anti-MS4A1 antibody [SP32] (ab64088) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab64088 was shown to bind specifically to MS4A1. 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. 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-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-CD20 antibody [SP32] (ab64088) at 1/1000 dilution

Lane 1:

Wild-type Raji cell lysate

Lane 2:

MS4A1 knockout Raji cell lysate

Lane 2:

Western blot - Human MS4A1 (CD20) knockout Raji cell line (<a href='/products/cell-lines/human-ms4a1-cd20-knockout-raji-cell-line-ab273871'>ab273871</a>)

Lane 3:

Ramos cell lysate

Lane 4:

A549 cell lysate

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

false