CD20 Recombinant Rabbit Monoclonal Antibody [PD00-02]
cat.: HA721138
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Mouse
Applications: IHC-P, mIHC, WB, IF-Cell, IP
Clonality: Monoclonal
Clone number: PD00-02
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer: PBS (pH7.4), 0.1% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Purification: Protein A affinity purified.
Molecular weight: Predicted band size: 33 kDa
Isotype: IgG
Immunogen: Synthetic peptide within Human CD20 aa 210-297 (Cytoplasmic).
Positive control: Human tonsil tissue, human lymph nodes tissue, human spleen tissue, human small cell lung cancer, human non-small cell lung cancer, human prostate cancer, human non-small cell lung cancer, mouse spleen tissue, Raji cell lysate, Ramos cell lysate, Daudi cell lysate.
Subcellular location: Cell membrane.
Recommended Dilutions:
  IHC-P
  mIHC
  WB
  IF-Cell
  IP
1:1,000-1:2,000
1:1,500-1:3,000
1:2,000
1:500
1-2μg/sample
Uniprot #: SwissProt: P11836 Human | P19437 Mouse
Alternative names: APY ATOPY B lymphocyte antigen CD20 B Lymphocyte Cell Surface Antigen B1 B-lymphocyte antigen CD20 B-lymphocyte cell-surface antigen B1 B-lymphocyte surface antigen B1 B1 Bp 35 Bp35 CD 20 CD20 CD20 antigen CD20 receptor CD20_HUMAN CVID 5 CVID5 Fc epsilon receptor I beta chain Fc Fragment of IgE high affinity I receptor for beta polypeptide FCER1B High affinity immunoglobulin epsilon receptor subunit beta IgE Fc receptor subunit beta IGEL IGER IGHER LEU 16 Leu-16 LEU16 leukocyte surface antigen Leu 16 Leukocyte surface antigen Leu-16 Ly44 Membrane spanning 4 domains A1 Membrane spanning 4 domains subfamily A member 2 membrane-spanning 4-domains A1 Membrane-spanning 4-domains subfamily A member 1 MGC3969 MS4A1 MS4A2 S7
Images
HA721138_1.jpg Fig1: Fluorescence multiplex immunohistochemical analysis of Tertiary Lymphoid Structures in Human Small Cell Lung Cancer (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-CD20 (HA721138, green), anti-PD-L1 (HA721176, cyan), anti-CD56 (ET1702-43, magenta) and anti-CD3 (HA720082, yellow) on tertiary lymphoid structures. Panel B: anti- CD20 stained on B cells. Panel C: anti-PD-L1 stained on dendritic cells and macrophages cells. Panel D: anti-CD56 stained on NKT cells. Panel E: anti-CD3 stained on T cells. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in four rounds of staining: in the order of HA721138 (1/1,500 dilution), HA721176 (1/1,000 dilution), ET1702-43 (1/1,000 dilution), and HA720082 (1/500 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_2.jpg Fig2: Fluorescence multiplex immunohistochemical analysis of the human non-small cell lung cancer (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-CD20 (HA721138, green), anti-CD68 (HA601115, gray), anti-PD-L1 (HA721176, cyan), anti-panCK (HA601138, magenta) and anti-CD3 (HA720082, yellow) on human non-small cell lung cancer. Panel B: anti- CD20 stained on B cells. Panel C: anti-CD68 stained on macrophage M1 and macrophage M2. Panel D: anti-PD-L1 stained on dendritic cells and macrophages cells. Panel E: anti-panCK stained on cancer cells. Panel F: anti-CD3 stained on T cells. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in five rounds of staining: in the order of HA721138 (1/1,500 dilution), HA601115 (1/2,000 dilution), HA721176 (1/1,000 dilution), HA601138 (1/3,000 dilution), and HA720082 (1/500 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_3.jpg Fig3: Fluorescence multiplex immunohistochemical analysis of tertiary lymphoid structures in human prostate cancer (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-CD20 (HA721138, green), anti-CD21 (HA721163, cyan) and anti-CD4 (ET1609-52, yellow) on tertiary lymphoid structures. Panel B: anti- CD20 stained on B cells. Panel C: anti-CD21 stained on naive B-cell, memory B-cell and plasma cells. Panel D: anti-CD4 stained on helper T cells and Treg cells. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in three rounds of staining: in the order of HA721138 (1/1,500 dilution), HA721163 (1/1,000 dilution), and ET1609-52 (1/1,000 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_4.jpg Fig4: Fluorescence multiplex immunohistochemical analysis of Human tonsil (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-BCL6 (HA601083, Red), anti-HLA-DPB1 (ET1704-13, Green), anti-Tryptase (ET1610-64, White), anti-CD20 (HA721138, Magenta) and anti-CD45 (ET7111-03, Yellow) on tonsil. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in five rounds of staining: in the order of HA601083 (1/200 dilution), ET1704-13 (1/2,000 dilution), ET1610-64 (1/5,000 dilution), HA721138 (1/2,000 dilution) and ET7111-03 (1/500 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_5.jpg Fig5: Fluorescence multiplex immunohistochemical analysis of Human tonsil (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-CD68 (HA601115, Red), anti-CD38 (HA721268, Green), anti-CD23 (HA721139, White), anti-CD11C (ET1606-19, Cyan), anti-CD45 (ET7111-03, Magenta) and anti-CD20 (HA721138, Yellow) on tonsil. Panel B: anti-CD68 stained on Macrophage. Panel C: anti-CD38 stained on lymphocyte subsets. Panel D: anti-CD11C stained on dendritic cells. Panel E: CD45 stained on lymphocytes. Panel F: anti-CD20 stained on B cells. Panel G: anti-CD23 stained on follicular dendritic cells. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in six rounds of staining: in the order of HA601115 (1/2,000 dilution), HA721268 (1/1,000 dilution), ET1606-19 (1/1,000 dilution), ET7111-03 (1/500 dilution), HA721138 (1/2,000 dilution) and HA721139 (1/800 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_6.jpg Fig6: Fluorescence multiplex immunohistochemical analysis of Human non-small cell lung cancer (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-PD-L1 (HA721176, red), anti-CD34 (ET1606-11, green), anti-Pan-CK (HA601138, cyan), anti-CD20 (HA721138, magenta), anti-αSMA (ET1607-53, yellow) and anti-CD57 (HA601114, white) on NSCLC. Panel B: anti-PD-L1 stained on dendritic cells and macrophages cells. Panel C: anti- CD34 stained on endothelial cells. Panel D: anti-Pan-CK stained on cancer cells. Panel E: CD20 stained on B cells. Panel F: anti-αSMA stained on cancer-associated fibroblasts and smooth muscle cells. Panel G: anti-CD57 stained on NK cells and T cells. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in six rounds of staining: in the order of HA721176 (1/1,000 dilution), ET1606-11 (1/1,000 dilution), HA601138 (1/3,000 dilution), HA721138 (1/2,000 dilution), ET1607-53 (1/3,000 dilution) and HA601114 (1/1,000 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner.
HA721138_7.jpg Fig7: Fluorescence multiplex immunohistochemical analysis of human tonsil (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-CD20 (HA721138, Cyan), anti-CD38 (HA721268, Violet) and anti-CD57 (HA601114, Yellow) on tonsil. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in three rounds of staining: in the order of HA721138 (1/2,000 dilution), HA721268 (1/1,000 dilution) and HA601114 (1/1,000 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Zeiss Observer 7 Inverted Fluorescence Microscope.
HA721138_8.jpg Fig8: Western blot analysis of CD20 on different lysates with Rabbit anti-CD20 antibody (HA721138) at 1/2,000 dilution.

Lane 1: Raji cell lysate
Lane 2: 293T cell lysate (negative)
Lane 3: Ramos cell lysate
Lane 4: HeLa cell lysate (negative)
Lane 5: Daudi cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 33 kDa
Observed band size: 33 kDa

Exposure time: 4 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721138) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HA721138_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-CD20 antibody (HA721138) at 1/2,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA721138) at 1/2,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HA721138_10.jpg Fig10: Immunohistochemical analysis of paraffin-embedded human lymph nodes tissue with Rabbit anti-CD20 antibody (HA721138) at 1/2,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA721138) at 1/2,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HA721138_11.jpg Fig11: Immunohistochemical analysis of paraffin-embedded human spleen tissue with Rabbit anti-CD20 antibody (HA721138) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA721138) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HA721138_12.jpg Fig12: Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-CD20 antibody (HA721138) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA721138) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HA721138_13.jpg Fig13: Immunocytochemistry analysis of Raji (positive) and 293T (negative) labeling CD20 with Rabbit anti-CD20 antibody (HA721138) at 1/500 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-CD20 antibody (HA721138) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HA721138_14.jpg Fig14: CD20 was immunoprecipitated from 0.2 mg Raji cell lysate with HA721138 at 2 µg/25 µl agarose. Western blot was performed from the immunoprecipitate using HA721138 at 1/2,000 dilution. Mouse anti Rabbit IgG heavy chain (Fc) secondary antibody (M1003-7) at 1/10,000 dilution was used for 1 hour at room temperature.

Lane 1: Raji cell lysate (input)
Lane 2: HA721138 IP in Raji cell lysate
Lane 3: Rabbit IgG instead of HA721138 in Raji cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 59 seconds; ECL: K1801
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.