Recombinant Anti-CD44 antibody [Hermes-3] - BSA and Azide free (ab255946)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Mouse monoclonal [Hermes-3] to CD44 - BSA and Azide free
- Suitable for: ICC/IF, IP, ELISA, IHC-P, WB
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-CD44 antibody [Hermes-3] - BSA and Azide free
See all CD44 primary antibodies -
Description
Mouse monoclonal [Hermes-3] to CD44 - BSA and Azide free -
Host species
Mouse -
Tested applications
Suitable for: ICC/IF, IP, ELISA, IHC-P, WBmore details -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant fragment corresponding to Human CD44.
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Positive control
- WB: HAP1, A549 and HeLa whole cell lysate. IHC-P: Human skin tissue. Human bladder carcinoma tissue. IP: HAP1 cell lysate. ICC/IF: HAP1, wild-type HeLa cells.
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General notes
ab255946 is the carrier-free version of ab254530.
This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
Hermes-3 -
Isotype
IgG2a -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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KO cell lines
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab255946 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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ICC/IF |
1/500.
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IP |
Use at an assay dependent concentration.
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ELISA |
Use 100-1000µg for 10 cells.
100-1000 ng/ml. |
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IHC-P | (1) |
Use a concentration of 0.107 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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WB |
Use a concentration of 0.536 µg/ml. Predicted molecular weight: 81 kDa.
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Notes |
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ICC/IF
1/500. |
IP
Use at an assay dependent concentration. |
ELISA
Use 100-1000µg for 10 cells. 100-1000 ng/ml. |
IHC-P
Use a concentration of 0.107 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
WB
Use a concentration of 0.536 µg/ml. Predicted molecular weight: 81 kDa. |
Target
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Function
Receptor for hyaluronic acid (HA). Mediates cell-cell and cell-matrix interactions through its affinity for HA, and possibly also through its affinity for other ligands such as osteopontin, collagens, and matrix metalloproteinases (MMPs). Adhesion with HA plays an important role in cell migration, tumor growth and progression. Also involved in lymphocyte activation, recirculation and homing, and in hematopoiesis. Altered expression or dysfunction causes numerous pathogenic phenotypes. Great protein heterogeneity due to numerous alternative splicing and post-translational modification events. -
Tissue specificity
Isoform 10 (epithelial isoform) is expressed by cells of epithelium and highly expressed by carcinomas. Expression is repressed in neuroblastoma cells. -
Sequence similarities
Contains 1 Link domain. -
Domain
The lectin-like LINK domain is responsible for hyaluronan binding. -
Post-translational
modificationsProteolytically cleaved in the extracellular matrix by specific proteinases (possibly MMPs) in several cell lines and tumors.
N-glycosylated.
O-glycosylated; contains more-or-less-sulfated chondroitin sulfate glycans, whose number may affect the accessibility of specific proteinases to their cleavage site(s).
Phosphorylated; activation of PKC results in the dephosphorylation of Ser-706 (constitutive phosphorylation site), and the phosphorylation of Ser-672. -
Cellular localization
Membrane. - Information by UniProt
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Database links
- Entrez Gene: 960 Human
- Omim: 107269 Human
- SwissProt: P16070 Human
- Unigene: 502328 Human
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Alternative names
- LHR antibody
- BA-1 antibody
- CD 44 antibody
see all
Images
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue staining CD44 with ab255946 at 1/200 dilution and co-stained with ab256584. Secondary antibody used was Alexa Fluor® 488 Donkey anti-mouse IgG (H+L)at 1/200 dilution. The tissue was incubated for 18 hours with PBS +2% normal human serum at 4°C. Blocking was done with 5% serum for 1 hour at room temperature. Heat mediated antigen retrieval with Tris-EDTA 1mM PH9.
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This data was developed using the same antibody clone in a different buffer formulation (ab254530). ab254530 staining CD44 in wild-type HeLa cells (top panel) and CD44 knockout HeLa cells (ab262515)(bottom panel). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Tween 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 ab254530 at 0.4μg/ml concentration and ab6046 (Rabbit polyclonal to beta 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 mouse IgG (Alexa Fluor® 488) (ab150117) at 2 μg/ml (shown in green) and a goat secondary antibody to rabbit IgG (Alexa Fluor® 594) (ab150080) 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). -
All lanes : Anti-CD44 antibody [Hermes-3] (ab254530) at 1/1000 dilution
Lane 1 : Wild-type HAP1 cell lysate
Lane 2 : CD44 knockout HAP1 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 81 kDaThis data was developed using the same antibody in a different buffer formulation (ab254530).
ab254530 was shown to react with CD44 in wild-type HAP1 cells in Western blot with loss of signal observed in a CD44 knockout cell line. Wild-type HAP1 and CD44 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab254530 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with goat anti-mouse HRP secondary antibodies at 0.3ug/mL before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, Glycerol and sodium azide (ab254530).
Immunohistochemical analysis of paraffin-embedded human skin tissue labeling CD44 with ab254530 at 0.107µg/ml, followed by ready to use secondary. Membranous staining on human skin is observed. The section was incubated with ab254530 for 5 mins at RT. The immunostaining staining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, followed by ready to use secondary.Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.
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This data was developed using the same antibody clone in a different buffer formulation (ab254530). ab254530 was shown to react with CD44 in wild-type HAP1 cells in Immunocytochemistry with loss of signal observed in a CD44 knockout cell line. Wild-type and knockout cells were mixed and pelleted at a 1:1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1/2000. The cells were then incubated with ab254530 at 1/500 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a secondary antibody to (Alexa Fluor® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880). These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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This data was developed using the same antibody clone in a different buffer formulation (ab255946). Immunoprecipitation of CD44 in HAP1 cells. Lysates were prepared and immunoprecipitation was performed using 1.0 μg of ab254530 pre-coupled to prot.G-Sepharose beads. Samples were washed and processed for western blot with ab189524 at 1/2000. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitate. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
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All lanes : Anti-CD44 antibody [Hermes-3] (ab254530) at 0.536 µg/ml
Lane 1 : A549 (human lung carcinoma epithelial cell) whole cell lysate
Lane 2 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Peroxidase-Conjugated Goat anti-Mouse IgG (H+L) at 1/10000 dilution
Predicted band size: 81 kDa
Exposure time: 1 secondThis data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, Glycerol and sodium azide (ab254530).
Blocking/Dilution buffer: 5% NFDM/TBST.
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, Glycerol and sodium azide (ab254530).
Immunohistochemical analysis of paraffin-embedded human bladder carcinoma tissue labeling CD44 with ab254530 at 0.107µg/ml, followed by ready to use secondary. Membranous staining on human bladder carcinoma is observed. The section was incubated with ab254530 for 5 mins at RT. The immunostaining staining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, followed by ready to use secondary.Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, Glycerol and sodium azide (ab254530).
ELISA - Anti-CD44 antibody [Hermes-3] (ab254530).
Antigen: Human CD44.
ab254530 used at 1000-0 ng/ml.
Secondary is an Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Mouse IgG (H+L) used at a 1/1000 dilution.
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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Datasheet download
References (0)
ab255946 has not yet been referenced specifically in any publications.