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  1. Link

    ccl4mip-1-beta-antibody-ep521y-ab45690.pdf

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Immunology Innate Immunity Macrophage / Inflamm.
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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)

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Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Immunocytochemistry/ Immunofluorescence - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Immunoprecipitation - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • ELISA - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
  • Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EP521Y] to CCL4/MIP-1 beta
  • Suitable for: ELISA, IP, WB, ICC/IF
  • Knockout validated
  • Reacts with: Mouse, Human

Conjugates logo Related conjugates and formulations

Carrier Free

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Human CCL4 knockout THP-1 cell line (ab273719)

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Overview

  • Product name

    Anti-CCL4/MIP-1 beta antibody [EP521Y]
    See all CCL4/MIP-1 beta primary antibodies
  • Description

    Rabbit monoclonal [EP521Y] to CCL4/MIP-1 beta
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ELISA, IP, WB, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide within Human CCL4/MIP-1 beta aa 1 to the C-terminus (N terminal). The exact sequence is proprietary.
    Database link: P13236

  • Positive control

    • THP-1, Raw264.7
  • 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
    For more information see here.

    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

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 49% PBS, 50% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EP521Y
  • Isotype

    IgG
  • Research areas

    • Immunology
    • Innate Immunity
    • Macrophage / Inflamm.
    • Immunology
    • Innate Immunity
    • Chemokines
    • Beta Chemokines (CC)
    • Kits/ Lysates/ Other
    • Kits
    • ELISA Kits
    • ELISA Kits
    • Cytokines and cytokine receptors ELISA kits
    • Immunology
    • Immune System Diseases
    • Antiviral Signaling
    • HIV-related
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Anti-CCL4/MIP-1 beta antibody [EP521Y] - BSA and Azide free (ab187674)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Recombinant Protein

    • Recombinant human CCL4/MIP-1 beta protein (ab9676)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab45690 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
ELISA
Use at an assay dependent concentration.
IP
1/60.
WB
1/1000 - 1/500000. Detects a band of approximately 10 kDa (predicted molecular weight: 10 kDa).
ICC/IF
1/100.
Notes
ELISA
Use at an assay dependent concentration.
IP
1/60.
WB
1/1000 - 1/500000. Detects a band of approximately 10 kDa (predicted molecular weight: 10 kDa).
ICC/IF
1/100.

Target

  • Function

    Monokine with inflammatory and chemokinetic properties. Binds to CCR5. One of the major HIV-suppressive factors produced by CD8+ T-cells. Recombinant MIP-1-beta induces a dose-dependent inhibition of different strains of HIV-1, HIV-2, and simian immunodeficiency virus (SIV). The processed form MIP-1-beta(3-69) retains the abilities to induce down-modulation of surface expression of the chemokine receptor CCR5 and to inhibit the CCR5-mediated entry of HIV-1 in T-cells. MIP-1-beta(3-69) is also a ligand for CCR1 and CCR2 isoform B.
  • Sequence similarities

    Belongs to the intercrine beta (chemokine CC) family.
  • Post-translational
    modifications

    N-terminal processed form MIP-1-beta(3-69) is produced by proteolytic cleavage after secretion from peripheral blood lymphocytes.
  • Cellular localization

    Secreted.
  • Target information above from: UniProt accession P13236 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 388372 Human
    • Entrez Gene: 6351 Human
    • Entrez Gene: 9560 Human
    • Entrez Gene: 20303 Mouse
    • Omim: 182284 Human
    • Omim: 603782 Human
    • Omim: 610757 Human
    • SwissProt: P13236 Human
    • SwissProt: Q8NHW4 Human
    • SwissProt: P14097 Mouse
    • Unigene: 655293 Human
    • Unigene: 661942 Human
    • Unigene: 75703 Human
    see all
  • Alternative names

    • MIP 1 beta antibody
    • Secreted protein G 26 antibody
    • ACT 2 antibody
    • ACT-2 antibody
    • ACT2 antibody
    • AT744.1 antibody
    • AT744.2 antibody
    • C C motif chemokine 4 antibody
    • C C motif chemokine 4 like antibody
    • C C motif chemokine ligand 4 like 1 antibody
    • C C motif chemokine ligand 4 like 2 antibody
    • CC chemokine ligand 4 antibody
    • CC chemokine ligand 4L1 antibody
    • CC chemokine ligand 4L1d2 antibody
    • CC chemokine ligand 4L2 antibody
    • CCL4 antibody
    • CCL4_HUMAN antibody
    • ccl4l 1 antibody
    • CCL4L antibody
    • CCL4L1 antibody
    • Chemokine (C C motif) ligand 4 antibody
    • Chemokine (C C motif) ligand 4 like 1 antibody
    • Chemokine (C C motif) ligand 4 like 1, telomeric antibody
    • Chemokine (C C motif) ligand 4 like 2 antibody
    • Chemokine CC Motif Ligand 4 antibody
    • G 26 antibody
    • G 26 T lymphocyte secreted protein antibody
    • G-26 T-lymphocyte-secreted protein antibody
    • HC21 antibody
    • Immune activation 2 antibody
    • LAG 1 antibody
    • LAG-1 antibody
    • LAG1 antibody
    • Lymphocyte activation gene 1 antibody
    • Lymphocyte activation gene 1 protein antibody
    • Macrophage inflammatory protein 1 beta antibody
    • Macrophage inflammatory protein 1-beta antibody
    • Macrophage inflammatory protein 1b2 antibody
    • MGC104418 antibody
    • MGC126025 antibody
    • MGC126026 antibody
    • MIP-1-beta antibody
    • MIP-1-beta(1-69) antibody
    • MIP-1-beta(3-69) antibody
    • MIP1 beta antibody
    • MIP1B antibody
    • MIP1B1 antibody
    • Monocyte adherence induced protein 5 alpha antibody
    • PAT 744 antibody
    • Protein H400 antibody
    • SCYA2 antibody
    • SCYA4 antibody
    • SCYA4L antibody
    • SCYA4L1 antibody
    • SCYA4L2 antibody
    • SCYQ4L2 antibody
    • Secreted protein G 26 antibody
    • Secreted protein G26 antibody
    • SIS gamma antibody
    • SIS-gamma antibody
    • Small inducible cytokine A4 (homologous to mouse Mip 1b) antibody
    • Small inducible cytokine A4 antibody
    • small inducible cytokine A4-like antibody
    • Small-inducible cytokine A4 antibody
    • T cell activation protein 2 antibody
    • T-cell activation protein 2 antibody
    see all

Images

  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    All lanes : Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690) at 1/1000 dilution

    Lane 1 : Wild-type THP-1 Vehicle control + Brefeldin A (5 u/mL, 6 h) cell lysate
    Lane 2 : Wild-type THP-1 Treated PMA (100 ng/mL, 56 h) + LPS (1 u/mL, 24 h) + Brefeldin A (5 u/mL, 6 h) cell lysate
    Lane 3 : CCL4 knockout THP-1 Vehicle control + Brefeldin A (5 u/mL, 6 h) cell lysate
    Lane 4 : CCL4 knockout THP-1 Treated PMA (100 ng/mL, 56 h) + LPS (1 u/mL, 24 h) + Brefeldin A (5 u/mL, 6 h) cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 10 kDa
    Observed band size: 12 kDa why is the actual band size different from the predicted?



    False colour image of Western blot: Anti-CCL4/MIP-1 beta antibody [EP521Y] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab45690 was shown to bind specifically to CCL4/MIP-1 beta. A band was observed at 12 kDa in wild-type THP-1 cell lysates with no signal observed at this size in CCL4 knockout cell line ab273719 (knockout cell lysate ab275512). To generate this image, wild-type and CCL4 knockout THP-1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    All lanes : Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690) at 1/1000 dilution

    Lane 1 : Untagged human CCL4 recombinant protein (aa24-92)
    Lane 2 : Untagged human CCL4L recombinant protein (aa24-92)
    Lane 3 : GST-tagged human CCL3 recombinant protein (aa27-92)
    Lane 4 : GST-tagged human CCL3L recombinant protein 2*(aa28-93)

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 10 kDa
    Observed band size: 12 kDa why is the actual band size different from the predicted?


    Exposure time: 5 seconds


    Blocking/Diluting buffer and concentration 5% NFDM /TBST 

  • Immunocytochemistry/ Immunofluorescence - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Immunocytochemistry/ Immunofluorescence - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)

    Immunocytochemistry/Immunofluorescence analysis of THP-1 (Human monocytic leukemia cell line) cells labeling CCL4/MIP-1 beta + CCL4L with ab45690 at 1/100. Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Cells were counter-stained with ab7291, ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution. DAPI was used to stain nuclei blue.

    The expression increased after treatment with Lipopolysaccharides (LPS), 100 ng/mL for 4 hours, followed by addition of Brefeldin A (1 μg/mL) for 3 hours.

  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    All lanes : Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690) at 1/1000 dilution

    Lane 1 : Untreated THP-1 (human acute monocytic leukemia) lysate
    Lane 2 : THP-1 treated with 100 nM Phorbol-12-myristate-13-acetate(PMA) overnight, then treated with Lipopolysaccharides (LPS) 100 ng/mL for 7 hours and then 1 µg/mL Brefeldin A was added for the last 3 hours, lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051)

    Predicted band size: 10 kDa
    Observed band size: 12 kDa why is the actual band size different from the predicted?


    Exposure time: 3 minutes


    Blocking/Diluting buffer and concentration 5% NFDM /TBST.

    CCL4/MIP-1 beta is induced in macrophages following exposure to bacterial LPS (PMID: 9848081).

  • Immunoprecipitation - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Immunoprecipitation - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)

    ab45690 at 1/60 immunoprecipitating CCL4/MIP-1 beta + CCL4L in THP-1 (Human monocytic leukemia cell line) whole cell lysate observed at 12 KDa (lanes 1 and 2).

    Lane 1 (input): THP-1 treated with 100 nM PMA overnight, then treated with 100 ng/mL LPS for 7 hours and 1 μg/mL Brefeldin A was added for the last 3 hours whole cell lysate, 10μg

    Lane 2 (+): ab45690 + THP-1 treated with 100 nM PMA overnight, then treated with 100 ng/mL LPS for 7 hours and 1 μg/mL Brefeldin A was added for the last 3 hours whole cell lysate

    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab45690 in THP-1 treated with 100 nM PMA overnight, then treated with 100 ng/mL LPS for 7 hours and 1 μg/mL Brefeldin A was added for the last 3 hours whole cell lysate

    For western blotting, ab45690 at 1/1000 and ab131366 VeriBlot for IP Detection Reagent (HRP) was used for detection (1/1000).

    Blocking/Diluting buffer 5% NFDM/TBST

  • Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Western blot - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    All lanes : Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690) at 1/1000 dilution

    Lane 1 : Untreated Raw264.7 (mouse abelson murine leukemia virus-induced tumor) whole cell lysate
    Lane 2 : Raw264.7 (mouse abelson murine leukemia virus-induced tumor) treated with LPS 10µg/mL for 4 hours and then 1 µg/mL Brefeldin A was added for the last 3 hours lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 10 kDa


    Exposure time: 3 minutes


    Blocking/Diluting buffer and concentration 5% NFDM /TBST 

  • ELISA - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    ELISA - Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    ELISA analysis of Human CCL4/MIP-1 beta recombinant protein at 500 ng/mL with ab45690. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as the secondary antibody.
  • Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)
    Anti-CCL4/MIP-1 beta antibody [EP521Y] (ab45690)

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.

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (7)

Publishing research using ab45690? Please let us know so that we can cite the reference in this datasheet.

ab45690 has been referenced in 7 publications.

  • Yang L  et al. Immune Profiling Reveals Molecular Classification and Characteristic in Urothelial Bladder Cancer. Front Cell Dev Biol 9:596484 (2021). PubMed: 33777927
  • Zhao H & He Y The Inhibitory Effect of Lysophosphatidylcholine on Proangiogenesis of Human CD34+ Cells Derived Endothelial Progenitor Cells. Front Mol Biosci 8:682367 (2021). PubMed: 34179086
  • Gao M  et al. Therapy With Carboplatin and Anti-PD-1 Antibodies Before Surgery Demonstrates Sustainable Anti-Tumor Effects for Secondary Cancers in Mice With Triple-Negative Breast Cancer. Front Immunol 11:366 (2020). PubMed: 32194569
  • Ren F  et al. Transcriptome analysis reveals GPNMB as a potential therapeutic target for gastric cancer. J Cell Physiol 235:2738-2752 (2020). PubMed: 31498430
  • Ren C  et al. An effective and biocompatible polyethylenimine based vaginal suppository for gene delivery. Nanomedicine 20:101994 (2019). PubMed: 31028885
  • Chen P  et al. Identification and validation of four hub genes involved in the plaque deterioration of atherosclerosis. Aging (Albany NY) 11:6469-6489 (2019). PubMed: 31449494
  • Zsiros E  et al. The Ovarian Cancer Chemokine Landscape Is Conducive to Homing of Vaccine-Primed and CD3/CD28-Costimulated T Cells Prepared for Adoptive Therapy. Clin Cancer Res 21:2840-50 (2015). PubMed: 25712684

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