Anti-CD64 antibody [EPR26480-569] - BSA and Azide free

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded (A) HEK-293T (human epithelial cell line from embryonic kidney) transfected with a Fcgr1 expression vector containing a Myc-His tag. (B) HEK-293T transfected with empty vector containing a Myc-His tag. tissue labeling CD64 with ab323507 at 1/2000 (0.241 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) HEK-293T (human epithelial cell line from embryonic kidney) transfected with a Fcgr1 expression vector containing a Myc-His tag negative staining on (B) HEK-293T transfected with empty vector containing a Myc-His tag. The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) (Right) / C2C12 (mouse myoblast) (Left) cells labelling CD64 with ab323507 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

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

Gated on viable cells.
Negative control : C2C12.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of Mouse bone marrow cells labelling CD64 with ab323507 at 1/500 dilution (0.1ug) / Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat anti-Rabbit IgG (Alexa Fluor® 647 ab150083) at 1/5000 dilution was used as the secondary antibody.

Gated on viable cells.
Cells were co-stained with anti mouse CD3 conjugated to Alexa Fluor®488.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of Mouse bone marrow cells labelling CD64 with ab323507 at 1/500 dilution (0.1ug) / Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat anti-Rabbit IgG (Alexa Fluor® 647 ab150083) at 1/5000 dilution was used as the secondary antibody.

Gated on viable cells.
Cells were co-stained with anti mouse CD11b conjugated to Brilliant Violet 421.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded (A) RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cell pellets (B) C2C12 (mouse myoblast) cell pellet tissue labeling CD64 with ab323507 at 1/100 (4.81 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) RAW264.7 cell pellet no staining on (B) C2C12 cell pellet. The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse cardiac muscle tissue labeling CD64 with ab323507 at 1/100 (4.81 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : positive staining on scattered immune cells of mouse cardiac muscle. The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling CD64 with ab323507 at 1/100 (4.81 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on glial cells of mouse cerebrum (PMID : 23452918). The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling CD64 with ab323507 at 1/100 (4.81 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse spleen (PMID : 26512139). The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling CD64 with ab323507 at 1/100 (4.81 ug/ml) dilution followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on Kupffer cells of mouse liver. The section was incubated with ab323507 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 LeicaDS9800 (Bond™ Polymer Refine Detection).

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed 0.1% TritonX-100 permeabilized J774A.1 (mouse reticulum cell sarcoma monocyte/macrophage) cells labelling CD64 with ab323507 at 1/50 (9.62 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing mambrane and weak cytoplasmic staining in J774A.1 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Negative control : NIH/3T3.
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 dilution (Magenta). 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 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed 0.1% TritonX-100 permeabilized RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cells labelling CD64 with ab323507 at 1/50 (9.62 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing mambrane and weak cytoplasmic staining in RAW 264.7 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Negative control : C2C12.
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 dilution (Magenta). 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 dilution.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of J774A.1 (mouse reticulum cell sarcoma monocyte/macrophage) (Right) / NIH/3T3 (mouse embryonic fibroblast) (Left) cells labelling CD64 with ab323507 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

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

Gated on viable cells.
Negative control : NIH/3T3.

• WB

Supplier Data

Western blot - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

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

Low expression : heart

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

All lanes:

Western blot - Anti-CD64 antibody [EPR26480-569] (<a href='/en-us/products/primary-antibodies/cd64-antibody-epr26480-569-ab323507'>ab323507</a>) at 1/1000 dilution

Lane 1:

Mouse bone marrow at 20 µg

Lane 2:

Mouse spleen tissue lysate at 20 µg

Lane 3:

Mouse heart tissue lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Observed band size: 60-75 kDa

false

Exposure time: 147s

• WB

Supplier Data

Western blot - Anti-CD64 antibody [EPR26480-569] - BSA and Azide free (AB325955)

This data was developed using ab323507, the same antibody clone in a different buffer formulation.

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

Negative control : C2C12 NIH/3T3 EL4 A20

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 21124939).

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

All lanes:

Western blot - Anti-CD64 antibody [EPR26480-569] (<a href='/en-us/products/primary-antibodies/cd64-antibody-epr26480-569-ab323507'>ab323507</a>) at 1/1000 dilution

Lane 1:

RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 2:

J774A.1 (mouse reticulum cell sarcoma monocyte/macrophage ) whole cell lysate at 20 µg

Lane 3:

C2C12 (mouse myoblast) whole cell lysate at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 5:

EL4 (mouse lymphoma T lymphocyte) whole cell lysate at 20 µg

Lane 6:

A20 (mouse reticulum sarcoma B lymphocyte) 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 1/100000 dilution

Observed band size: 60-75 kDa,36 kDa

false

Exposure time: 180s