Anti-CD204 抗体 [RM2033]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CD204 antibody [RM2033] (AB315834)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized THP-1 (human monocytic leukemia monocyte) cells labelling CD204 with ab315834 at 1/100 (5.14 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 cytoplasmic staining in THP-1 cells treated with Phorbol-12-myristate-13-acetate (80nM) for 72hours.
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 (2.5ug/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-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling CD204 with ab315834 at 1/500 (1.028 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on kupffer cells of human liver. The section was incubated with ab315834 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

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Human lung tissue labeling CD204 with ab315834 at 1/500 (1.028 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on macrophages of human lung. The section was incubated with ab315834 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.

• IP

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Immunoprecipitation - Anti-CD204 antibody [RM2033] (AB315834)

CD204 was immunoprecipitated from 0.35 mg THP-1 (human monocytic leukemia monocyte) treated with 80nM Phorbol-12-myristate-13-acetate (PMA) for 72 hours, whole cell lysate with ab315834 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315834 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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

All lanes:

Immunoprecipitation - Anti-CD204 antibody [RM2033] (ab315834) at 1/1000 dilution

Lane 1:

THP-1 (human monocytic leukemia monocyte) treated with 80nM Phorbol-12-myristate-13-acetate (PMA) for 72 hours, whole cell lysate

Lane 2:

ab315834 at 1/30 IP in THP-1 (human monocytic leukemia monocyte) treated with 80nM Phorbol-12-myristate-13-acetate (PMA) for 72 hours, whole cell lysate at 1/30 dilution

Lane 3:

Rabbit monoclonal IgG (<a href='/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab315834 in THP-1 treated with 80nM Phorbol-12-myristate-13-acetate (PMA) for 72 hours, whole cell lysate

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

false

Exposure time: 10s

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Mouse lung tissue labeling CD204 with ab315834 at 1/2000 (0.257 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on macrophages of mouse lung. The section was incubated with ab315834 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

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Immunocytochemistry/ Immunofluorescence - Anti-CD204 antibody [RM2033] (AB315834)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse bone marrow cells labelling CD204 with ab315834 at 1/100 (5.14 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 increased cytoplasmic staining in mouse bone marrow cells treated with Lipopolysaccharides (2ug/ml) for 24h, which is co-staining with F4/80.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-F4/80 rat monoclonal antibody was used to counterstain at 1/200 (2.5ug/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-CD204 antibody [RM2033] (AB315834)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cells labelling CD204 with ab315834 at 1/100 (5.14 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 cytoplasmic staining in RAW 264.7 cells.
Negative control : Neuro-2a.
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 (2.5ug/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.

• Flow Cyt

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Flow Cytometry - Anti-CD204 antibody [RM2033] (AB315834)

Flow cytometric analysis of Isotype (Left) / Mouse bone marrow treated with 2ug/ml LPS for 24h (Middle) / Untreated mouse bone marrow (Right) cells labelling CD204 with ab315834 at 1/5000 dilution (0.01 ug)/Middle and 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 F4/80 conjugated to Allophycocyanin.

• Flow Cyt

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Flow Cytometry - Anti-CD204 antibody [RM2033] (AB315834)

Flow cytometric analysis of Neuro-2a (mouse neuroblastoma neuroblast, Left) / Raw264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage, Right) cells labelling CD204 with ab315834 at 1/50 dilution (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 : Neuro-2a. Gated on viable cells.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Mouse head and neck Neuroendocrine carcinoma tissue labeling CD204 with ab315834 at 1/2000 (0.257 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse head and neck Neuroendocrine carcinoma. The section was incubated with ab315834 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-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded (A) RAW 264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) cell pellets. (B) Neuro-2a (mouse neuroblastoma neuroblast) cell pellets. tissue labeling CD204 with ab315834 at 1/2000 (0.257 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) RAW 264.7 cell pellets, no staining on (B) Neuro-2a cell pellets. The section was incubated with ab315834 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-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Mouse colon tissue labeling CD204 with ab315834 at 1/2000 (0.257 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on immune cells of mouse colon. The section was incubated with ab315834 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-CD204 antibody [RM2033] (AB315834)

Immunohistochemical analysis of paraffin-embedded Mouse Burkitt's lymphoma tissue labeling CD204 with ab315834 at 1/2000 (0.257 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse Burkitt's lymphoma. The section was incubated with ab315834 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.

• WB

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Western blot - Anti-CD204 antibody [RM2033] (AB315834)

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

Negative control : Neuro-2a, NIH/3T3.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 35937296, 30991038).

CD204 expression can be induced by PMA treatment (PMID : 30991038).

This blot of lane 7 was developed using a higher sensitivity ECL substrate.

The identity of the higher MW bands at approximately 150 kDa in lane 1 and lane 7 are unknown.

Lanes 1-6 are applied with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 and lane 7 is applied with Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000.

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

Exposure time : Lane 1-6 : 37 seconds; Lane 7 : 180 seconds.

All lanes:

Western blot - Anti-CD204 antibody [RM2033] (ab315834) at 1/1000 dilution

Lane 1:

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

Lane 2:

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

Lane 3:

Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate at 20 µg

Lane 4:

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

Lane 5:

Untreated THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Lane 6:

THP-1 treated with 80nM Phorbol-12-myristate-13-acetate (PMA) for 72 hours, whole cell lysate at 20 µg

Lane 7:

Human liver tissue lysate at 20 µg

Secondary

Lanes 1 - 6:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Lane 7:

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

Observed band size: 80 kDa,36 kDa

false