Recombinant Anti-Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 antibody [EPR23751-54] (ab259372)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23751-54] to MIP-1 alpha/CCL3 + CCL3L1
- Suitable for: Flow Cyt (Intra), WB, IHC-P, IP, ICC/IF
- Reacts with: Mouse, Human
Related conjugates and formulations
Overview
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Product name
Anti-Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 antibody [EPR23751-54] -
Description
Rabbit monoclonal [EPR23751-54] to MIP-1 alpha/CCL3 + CCL3L1 -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), WB, IHC-P, IP, ICC/IFmore details -
Species reactivity
Reacts with: Mouse, Human -
Immunogen
This product was produced with the following immunogens:
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers. -
Positive control
- WB: THP-1 (treated with PMA / LPS(Lipopolysaccharide / Brefeldin A) whole cell lysate; RAW264.7 (treated with LPS / Brefeldin A) whole cell lysate; Recombinant himan CCL3 protein; Recombinant human CCl3L1 protein. IHC-P: Human Hodgkin lymphoma tissue. ICC/IF: THP-1 and RAW264.7 cells (treated with PMA / LPS / Brefeldin A). Flow Cyt (intra): THP-1 and RAW264.7 cells (treated with PMA / LPS / Brefeldin A). IP: THP-1 (treated with PMA / LPS(Lipopolysaccharide / Brefeldin A) whole cell lysate.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23751-54 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab259372 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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Flow Cyt (Intra) |
1/500.
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WB |
1/1000. Detects a band of approximately 12 kDa (predicted molecular weight: 10 kDa).
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IHC-P |
1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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IP |
1/30.
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ICC/IF |
1/50.
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Notes |
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Flow Cyt (Intra)
1/500. |
WB
1/1000. Detects a band of approximately 12 kDa (predicted molecular weight: 10 kDa). |
IHC-P
1/1000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
IP
1/30. |
ICC/IF
1/50. |
Target
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Cellular localization
Macrophage Inflammatory Protein 1 alpha / CCL3: Secreted. CCL3L1: Secreted. -
Database links
- Entrez Gene: 414062 Human
- Entrez Gene: 6348 Human
- Entrez Gene: 6349 Human
- Entrez Gene: 20302 Mouse
- Omim: 182283 Human
- Omim: 609468 Human
- SwissProt: P10147 Human
- SwissProt: P16619 Human
see all
Images
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Immunohistochemical analysis of paraffin-embedded human Hodgkin lymphoma tissue labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human hodgkin Lymphoma (PMID: 31581676). The section was incubated with ab259372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counter stained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X100 permeabilized THP-1 (human monocytic leukemia monocyte) cells labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/100 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) secondary antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in THP-1 cells treated with 12-myristate 13-acetate (100 nM) for 16 hours then with lipopolysaccharide (100 ng/ml) for 4 hours and with brefeldin A (1μg/ml) for another 3 hours. Tubulin is counterstained using ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution (Red). The nuclear counterstain is DAPI (Blue).
Secondary antibody only control: Used PBS instead of priary antibody, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) secondary antibody at 1/1000 dilution.
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Intracellular flow cytometric analysis of 5% paraformaldehyde-fixed, 90% methanol-permeabilized THP-1 (Human monocytic leukemia monocyte) (treated with 100nM phorbol 12-myristate 13-acetate (PMA) for 16 hours, then 100ng/ml lipopolysaccharide (LPS) for 4 hours, and add 1ug/ml Brefeldin A (BFA) for another 3 hours) (Red)/ Untreated control (Green) cells labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/500 dilution (Red / Green) compared with a Isotype control details (ab172730) (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.
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All lanes : Anti-Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 antibody [EPR23751-54] (ab259372) at 1/1000 dilution
Lane 1 : Untreated THP-1 (human monocytic leukemia monocyte) whole cell lysate
Lane 2 : THP-1 (treated with 100nM PMA for 16 hours. And then replace PMA with 100 ng/ml LPS (Lipopolysaccharide) for 4 hours, and then 1 µg/ml Brefeldin A was added for the last 3 hours), whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/50000 dilution
Predicted band size: 10 kDa
Observed band size: 12 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 8 seconds.
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Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 was immunoprecipitated from 0.35 mg THP-1 (human monocytic leukemia monocyte) (treated with 100nM PMA for 16 hours. And then replace PMA with 100 ng/ml LPS (Lipopolysaccharide) for 7 hours, 1 μg/ml BFA was added for the last 3 hours) whole cell lysate whole cell lysate with ab259372 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab259372 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: THP-1 (treated as above) whole cell lysate 3 ug
Lane 2: ab259372 IP in THP-1 (treated as above) whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab259372 in THP-1 (treated as above) whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 8 seconds.
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All lanes : Anti-Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 antibody [EPR23751-54] (ab259372) at 1/5000 dilution
Lane 1 : Untreated RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate
Lane 2 : RAW 264.7 (treated with 100 ng/ml LPS(Lipopolysaccharide) for 4 hours and then 1 µg/ml Brefeldin A was added for the last 3 hours), whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 10 kDa
Observed band size: 12 kDa why is the actual band size different from the predicted?Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 3 seconds.
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X100 permeabilized RAW264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) cells labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/100 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) secondary antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in RAW 264.7 cells treated with 12-myristate 13-acetate (100 nM) for 16 hours then with lipopolysaccharide (100 ng/ml) for 4 hours and with brefeldin A (1μg/ml) for another 3 hours. Tubulin is counterstained using ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution (Red). The nuclear counterstain is DAPI (Blue).
Secondary antibody only control: Used PBS instead of priary antibody, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) secondary antibody at 1/1000 dilution.
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Intracellular flow cytometric analysis of5% paraformaldehyde-fixed, 90% methanol-permeabilized RAW264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) (treated with 100ng/ml lipopolysaccharide (LPS) for 4 hours, and add Brefeldin A (BFA) for another 3 hours (Red)/ Untreated control (Green)) (Red)/ Untreated control (Green) cells labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372at 1/500 dilution (Red / Green) compared with a Isotype control details (ab172730) (Black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). Goat anti rabbit IgG (Alexa Fluor®488, ab150077), at 1/2000 dilution was used as the secondary antibody.
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Immunohistochemical analysis of paraffin-embedded THP-1 cells labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on THP-1 treated with 100nM PMA, LPS and Brefeldin treatment (image A), and negative staining on untreat THP-1 (image B). The section was incubated with ab259372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counter stained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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All lanes : Anti-Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 antibody [EPR23751-54] (ab259372) at 1/1000 dilution
Lane 1 : His-tagged mouse CCL3 recombinant protein (aa24-92) * 2 ,10 ng
Lane 2 : GST/His-tagged human CCL3L1 recombinant protein (aa24-93), 10 ng
Lane 3 : LIF/His-tagged human CCL4 recombinant protein (aa24-92)*2, 10 ng
Lane 4 : His-tagged human CCL18 recombinant protein (aa21-89),10 ng
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/50000 dilution
Predicted band size: 10 kDaBlocking and dilution buffer: 5% NFDM/TBST.
Exposire time: 10 seconds.
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Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Negative control: No staining on human tonsil. The section was incubated with ab259372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counter stained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
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Immunohistochemical analysis of paraffin-embedded human spleen tissue labeling Macrophage Inflammatory Protein 1 alpha / CCL3 + CCL3L1 with ab259372 at 1/1000 dilution, followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Negative control: No staining on human spleen tissue. The section was incubated with ab259372 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counter stained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (1)
ab259372 has been referenced in 1 publication.
- Andreuzzi E et al. Colorectal cancer development is affected by the ECM molecule EMILIN-2 hinging on macrophage polarization via the TLR-4/MyD88 pathway. J Exp Clin Cancer Res 41:60 (2022). PubMed: 35148799