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    cxcr5-antibody-epr23463-30-ab254415.pdf

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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-CXCR5 antibody [EPR23463-30] (ab254415)

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  • SDS
  • Certificate of Compliance
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Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Immunocytochemistry/ Immunofluorescence - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
  • Anti-CXCR5 antibody [EPR23463-30] (ab254415)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR23463-30] to CXCR5
  • Suitable for: ICC/IF, WB, Flow Cyt, IHC-P
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Alexa Fluor® 488 Carrier Free PE

You may also be interested in

Secondary
Product image
Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
Primary
Product image
Anti-Reg3a antibody (ab198824)
Knockout
Product image
Human CXCR5 knockout Raji cell line (ab273380)

View more associated products

Overview

  • Product name

    Anti-CXCR5 antibody [EPR23463-30]
    See all CXCR5 primary antibodies
  • Description

    Rabbit monoclonal [EPR23463-30] to CXCR5
  • Host species

    Rabbit
  • Specificity

    We observe only weak staining in human WB.

  • Tested applications

    Suitable for: ICC/IF, WB, Flow Cyt, IHC-Pmore details
    Unsuitable for: IP
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: Raji, Daudi, Neuro-2a, A20, Mouse spleen, mouse lymph node and rat lymph node, C6 lysates. IHC-P: Human tonsil tissue. Human diffuse large B- lymphoma tissue. Flow Cyt: Human peripheral blood mononuclear and Raji cells. ICC/IF: Daudi cells.
  • 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. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.2
    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR23463-30
  • Isotype

    IgG
  • Research areas

    • Cardiovascular
    • Blood
    • Other
    • Immunology
    • Adaptive Immunity
    • B Cells
    • CD
    • Immunology
    • Innate Immunity
    • Chemokines
    • Alpha Chemokine Rec. (CXCR)
    • Signal Transduction
    • Signaling Pathway
    • G Protein Signaling
    • GPCR
    • Microbiology
    • Organism
    • Virus
    • RNA Virus
    • ssRNA positive strand virus
    • SARS Coronavirus
    • Cancer
    • Signal transduction
    • G protein signaling
    • GPCR
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Anti-CXCR5 antibody [EPR23463-30] - BSA and Azide free (ab272936)
    • Alexa Fluor® 488 Anti-CXCR5 antibody [EPR23463-30] (ab275172)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • KO cell lines

    • Human CXCR5 knockout Raji cell line (ab273380)
  • KO cell lysates

    • Human CXCR5 knockout Raji cell lysate (ab275531)
  • Related Products

    • Rabbit specific IHC polymer detection kit HRP/DAB (ab209101)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab254415 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
ICC/IF
Use a concentration of 5 µg/ml.
WB
1/1000. Predicted molecular weight: 42 kDa.

We observe only weak staining in human WB.

Flow Cyt (1)
1/500.
IHC-P
1/5000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

We do not suggest this product for use in IHC with mouse or rat.

Notes
ICC/IF
Use a concentration of 5 µg/ml.
WB
1/1000. Predicted molecular weight: 42 kDa.

We observe only weak staining in human WB.

Flow Cyt
1/500.
IHC-P
1/5000. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

We do not suggest this product for use in IHC with mouse or rat.

Application notes
Is unsuitable for IP.

Target

  • Function

    Cytokine receptor that binds to B-lymphocyte chemoattractant (BLC). Involved in B-cell migration into B-cell follicles of spleen and Peyer patches but not into those of mesenteric or peripheral lymph nodes. May have a regulatory function in Burkitt lymphoma (BL) lymphomagenesis and/or B-cell differentiation.
  • Tissue specificity

    Expression in mature B-cells and Burkitt lymphoma cells.
  • Sequence similarities

    Belongs to the G-protein coupled receptor 1 family.
  • Cellular localization

    Cell membrane.
  • Target information above from: UniProt accession P32302 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 643 Human
    • Entrez Gene: 12145 Mouse
    • Entrez Gene: 29363 Rat
    • Omim: 601613 Human
    • SwissProt: P32302 Human
    • SwissProt: Q04683 Mouse
    • SwissProt: P34997 Rat
    • Unigene: 113916 Human
    • Unigene: 490637 Mouse
    • Unigene: 6246 Mouse
    • Unigene: 127121 Rat
    see all
  • Alternative names

    • BLR 1 antibody
    • BLR1 antibody
    • Burkitt lymphoma receptor 1 antibody
    • Burkitt lymphoma receptor 1 GTP binding protein antibody
    • Burkitt lymphoma receptor 1, GTP binding protein (chemokine (C-X-C motif) receptor 5) antibody
    • C X C chemokine receptor type 5 antibody
    • C-X-C chemokine receptor type 5 antibody
    • CD 185 antibody
    • CD185 antibody
    • CD185 antigen antibody
    • Chemokine (C-X-C motif) receptor 5 antibody
    • Chemokine C X C motif receptor 5 antibody
    • Chemokine CXC motif receptor 5 antibody
    • Chemokine receptor 5 antibody
    • CXC chemokine receptor type 5 antibody
    • CXC R5 antibody
    • CXC-R5 antibody
    • CXCR 5 antibody
    • CXCR-5 antibody
    • Cxcr5 antibody
    • CXCR5_HUMAN antibody
    • MDR 15 antibody
    • MDR-15 antibody
    • MDR15 antibody
    • MGC117347 antibody
    • Monocyte derived receptor 15 antibody
    • Monocyte-derived receptor 15 antibody
    see all

Images

  • Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)

    Flow cytometry overlay histogram showing wild-type Raji (green line) and CXCR5 knockout Raji cells (ab273380) stained with ab254415 (red line). The cells were incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serumto block FC receptors and non-specific protein-protein interaction followed by the antibody (ab254415) (1x106 in 100μl at 0.2 μg/ml) for 30 min at 4°C.

    The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 4°C.

    Isotype control antibody was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody (wild-type Raji - black line; CXCR5 knockout Raji - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

    Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling CXCR5 with ab254415 at 1/5000 dilution followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human tonsil (PMID: 12393412). The section was incubated with ab254415 for 10 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    All lanes : Anti-CXCR5 antibody [EPR23463-30] (ab254415) at 1/1000 dilution

    Lane 1 : Wild-type Raji cell lysate
    Lane 2 : CXCR5 knockout Raji cell lysate
    Lane 3 : Daudi cell lysate
    Lane 4 : Jurkat cell lysate

    Lysates/proteins at 30 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 42 kDa



    Lanes 1 - 4: Merged signal (red and green). Green - ab254415 observed at 60 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

    ab254415 was shown to react with CXCR5 in Raji wild-type cells in Western blot with loss of signal observed in CXCR5 knockout sample. Wild-type and CXCR5 knockout Raji cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab254415 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Immunocytochemistry/ Immunofluorescence - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    ab254415 staining CXCR5 in Daudi cells (top panel, positive control) and Jurkat cells (bottom panel, negative control). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab254415 at 5μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
    Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).
  • Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)

    Flow cytometric analysis of Raji (Human Burkitt's lymphoma B lymphocyte) cells labelling CXCR5 with ab254415 at 1/500 dilution (0.1µg) (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, ab150077) at 1/2000 dilution was used as the secondary antibody. Gated on viable cells.

  • Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Flow Cytometry - Anti-CXCR5 antibody [EPR23463-30] (ab254415)

    Flow cytometric analysis of human peripheral blood mononuclear cell (PBMC) cells labelling CXCR5 with ab254415 at 1/500 dilution (0.1µg) (Right) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Left). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary at a 1/2000 dilution. Cells were stained with rabbit IgG (Left) or ab254415 (Right), then stained with anti-CD19 conjugated to Alexa Fluor® 647.

    Gated on viable cells.

  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    All lanes : Anti-CXCR5 antibody [EPR23463-30] (ab254415) at 1/1000 dilution

    Lane 1 : Neuro-2a (mouse neuroblastoma neuroblast), whole cell lysate
    Lane 2 : A20 (mouse reticulum sarcoma B lymphocyte), whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution

    Predicted band size: 42 kDa
    Observed band size: 42 kDa



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

    Samples are non-boiled as boiling may cause protein aggregates. 

    The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 30553016).

    Exposure time: 20 seconds.

  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    All lanes : Anti-CXCR5 antibody [EPR23463-30] (ab254415) at 1/1000 dilution

    Lane 1 : Rat lymph node tissue lysate
    Lane 2 : C6 (rat glial tumor glial cell), whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution

    Predicted band size: 42 kDa
    Observed band size: 42 kDa



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

    This blot was developed using a higher sensitivity ECL substrate.

    Samples are non-boiled as boiling may cause protein aggregates.

    The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 30553016).

    Exposure time: 122 seconds.

  • Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Western blot - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    All lanes : Anti-CXCR5 antibody [EPR23463-30] (ab254415) at 1/1000 dilution

    Lane 1 : Mouse spleen tissue lysate
    Lane 2 : Mouse lymph node tissue lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution

    Predicted band size: 42 kDa
    Observed band size: 42 kDa



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

    This blot was developed using a higher sensitivity ECL substrate.

    Samples are non-boiled as boiling may cause protein aggregates.

    The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 30553016).

    Exposure time: 122 seconds.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CXCR5 antibody [EPR23463-30] (ab254415)

    Immunohistochemical analysis of paraffin-embedded human diffuse large B-cell lymphoma tissue labeling CXCR5 with ab254415 at 1/5000 dilution followed by ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human diffuse large B-cell lymphoma. The section was incubated with ab254415 for 10 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 ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins.

  • Anti-CXCR5 antibody [EPR23463-30] (ab254415)
    Anti-CXCR5 antibody [EPR23463-30] (ab254415)

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

Certificate of Compliance

To download a Certificate of Compliance, please enter your Lot number below:

References (0)

Publishing research using ab254415? Please let us know so that we can cite the reference in this datasheet.

ab254415 has not yet been referenced specifically in any publications.

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Flow Cytometry abreview for Anti-CXCR5 antibody [EPR23463-30]

Average
Abreviews
Abreviews
abreview image
Application
Flow Cytometry
Sample
Mouse Cell (Lung leukocyte cell preparation)
Permeabilization
No
Gating Strategy
CXCR5 expression was assessed on Lineage+ cells. Lineage markers include CD3, TCRs, CD19, B220, CD11c, CD11b, NK1.1, Gr1, Ter119)
Specification
Lung leukocyte cell preparation
Preparation
Cell harvesting/tissue preparation method: Spleen is mashed against a 100um cell strainer by using a plunger until the tissue is dissociated. Cell strainer is washed with FACS buffer and cells are centrifuged at 400rcf for 5min at 4C. Cells are lysated with amonium clorhide for 5min at RT. Cell are washed and centrifuged again. Cells are resuspended in FACS Buffer.
Sample buffer: FACS Buffer (PBS supplemented with 2%FBS)
Fixation
not fixed
Read More
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Monica Romera-Hernandez

Verified customer

Submitted Jul 27 2021

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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