Anti-CD21 antibody [EPR27369-9]

• Flow Cyt

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Flow Cytometry - Anti-CD21 antibody [EPR27369-9] (AB315160)

Flow cytometric analysis of Human PBMC (human peripheral blood mononuclear cell) cells labelling CD21 with ab315160 at 1/500 dilution (0.1 ug)/Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control.

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Gated on viable cells. Cells are co-stained with CD19 conjugated to Alexa Fluor®647.

• Flow Cyt

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Flow Cytometry - Anti-CD21 antibody [EPR27369-9] (AB315160)

Flow cytometric analysis of Human PBMC (human peripheral blood mononuclear cell) cells labelling CD21 with ab315160 at 1/500 dilution (0.1 ug)/Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Gated on viable cells. Cells are co-stained with CD56 conjugated to Brilliant Violet 421.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD21 antibody [EPR27369-9] (AB315160)

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling CD21 with ab315160 at 1/50 (10.56 ug/ml) dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Positive staining on germinal center of human tonsil. The section was incubated with ab315160 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD21 antibody [EPR27369-9] (AB315160)

Immunohistochemical analysis of paraffin-embedded Human hodgkin's lymphoma tissue labeling CD21 with ab315160 at 1/50 (10.56 ug/ml) dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Positive staining on human hodgkin's lymphoma. The section was incubated with ab315160 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CD21 antibody [EPR27369-9] (AB315160)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized Human PBMC (human primary peripheral blood mononuclear cell) cells labelling CD21 with ab315160 at 1/50 (10.56 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing membranous staining in subsets of human PBMCs. Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5 ug/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CD21 antibody [EPR27369-9] (AB315160)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized Raji (human Burkitt's lymphoma B lymphocyte) cells labelling CD21 with ab315160 at 1/50 (10.56 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing membranous and cytoplasmic staining in Raji cell line. Negative control : K-562 (PMID : 6983911).Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5 ug/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD21 antibody [EPR27369-9] (AB315160)

Immunohistochemical analysis of paraffin-embedded Human cardiac muscle tissue labeling CD21 with ab315160 at 1/50 (10.56 ug/ml) dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Negative control : no staining on human cardiac muscle. The section was incubated with ab315160 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• Flow Cyt

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Flow Cytometry - Anti-CD21 antibody [EPR27369-9] (AB315160)

Flow cytometric analysis of K-562 (human chronic myelogenous leukemia lymphoblast, Left) / Raji (human Burkitt's lymphoma B lymphocyte, Right) cells labelling CD21 with ab315160 at 1/500 dilution (0.1 ug)/Red compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Negative control : K-562. Gated on viable cells.

• mIHC

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Multiplex immunohistochemistry - Anti-CD21 antibody [EPR27369-9] (AB315160)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human tonsil tissue staining CD21 with ab315160 at a 1 : 50 (10.56 ug/ml) dilution, ab237772 anti-CD19 used at 1 : 5000 (0.21 ug/ml) dilution and ab237707 anti-CD3E used at a 1 : 500 (1.01 ug/ml) dilution.

Panel A : merged staining of anti-CD21 (green; Opal™690), anti-CD3E (grey; Opal™520) and anti-CD19 (red; Opal™570) on human tonsil.
Panel B : anti-CD21 staining mature B lymphocytes and T-lymphocytes of human tonsil.
Panel C : anti-CD19 staining B lymphocytes of human tonsil.
Panel D : anti-CD3E staining T lymphocytes of human tonsil.

The section was incubated in three rounds of staining : in the order of ab315160, ab237772, and ab237707 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.

• IP

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Immunoprecipitation - Anti-CD21 antibody [EPR27369-9] (AB315160)

CD21 was immunoprecipitated from 0.35 mg Raji (human Burkitt's lymphoma B lymphocyte) with ab315160 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315160 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate

Lane 2 : ab315160 IP in Raji (human Burkitt's lymphoma B lymphocyte)

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab315160 in Raji whole cell lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-CD21 antibody [EPR27369-9] (ab315160) at 1/30 dilution

All lanes:

Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate

Secondary

All lanes:

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

false

Exposure time: 41s

• WB

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Western blot - Anti-CD21 antibody [EPR27369-9] (AB315160)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : Human heart.

The identity of the lower MW bands at approximately 50kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Exposure time : Lane 1 : 15 seconds Lane 2 : 180 seconds

All lanes:

Western blot - Anti-CD21 antibody [EPR27369-9] (ab315160) at 1/1000 dilution

Lane 1:

Human tonsil tissue lysate at 20 µg

Lane 2:

Human heart tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 150 kDa,36 kDa

false

• WB

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Western blot - Anti-CD21 antibody [EPR27369-9] (AB315160)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : K-562 (PMID : 6983911).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Exposure time : Lane 1 : 48 seconds Lane 2 : 180 seconds

All lanes:

Western blot - Anti-CD21 antibody [EPR27369-9] (ab315160) at 1/1000 dilution

Lane 1:

Raji (human Burkitt's lymphoma B lymphocyte) whole cell lysate at 20 µg

Lane 2:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 150 kDa,36 kDa

false

• WB

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Western blot - Anti-CD21 antibody [EPR27369-9] (AB315160)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-CD21 antibody [EPR27369-9] (ab315160) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a myc-His-tag®, whole cell lysate at 10 µg

Lane 2:

293T cells transfected with a human CD21 expression vector containing a myc-His-tag®, whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 150 kDa

false

Exposure time: 3.25s