Anti-CD20 抗体 [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.

See Abreview

See Abreview

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.

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 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 (green; Opal™520) and anti-CD20 (gray; Opal™570) on rat spleen.
Panel B: anti-Sialoadhesin/CD169 stained on macrophages.
Panel C: anti-CD3 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.

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 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 (green; Opal™520) and anti-CD20 (gray; Opal™570) on rat spleen.
Panel B: anti-Sialoadhesin/CD169 stained on macrophages.
Panel C: anti-CD3 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.

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).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat spleen tissue sections labeling CD20 with ab64088 at 1/100 dilution (3.11 µg/ml). Heat mediated antigen retrieval with citrate buffer (pH 6.0, epitope retrieval solution 1) for 10mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Hematoxylin was used as a counterstain. Positive staining on rat spleen, performed on a Leica Biosystems BOND™ RX instrument.
The section was incubated with ab64088 for 30 mins at room temperature

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.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Mouse spleen tissue sections labeling CD20 with ab64088 at 1/100 dilution (3.11 µg/ml). Heat mediated antigen retrieval with citrate buffer (pH 6.0, epitope retrieval solution 1) for 10mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Hematoxylin was used as a counterstain. Positive staining on mouse spleen, performed on a Leica Biosystems BOND™ RX instrument.
The section was incubated with ab64088 for 30 mins at room temperature.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human tonsil tissue sections labeling CD20 with ab64088 at 1/100 dilution (3.11 µg/ml). Heat mediated antigen retrieval with citrate buffer (pH 6.0, epitope retrieval solution 1) for 10mins. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Hematoxylin was used as a counterstain. Positive staining on human tonsil, performed on a Leica Biosystems BOND™ RX instrument.
The section was incubated with ab64088 for 30 mins at room temperature.

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).

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