Recombinant Anti-S100A4 antibody [EPR14639(2)] (ab197896)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR14639(2)] to S100A4
- Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF, IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-S100A4 antibody [EPR14639(2)]
See all S100A4 primary antibodies -
Description
Rabbit monoclonal [EPR14639(2)] to S100A4 -
Host species
Rabbit -
Specificity
Based on sequence homologies, the antibody may cross-react with other proteins of the same family (S100A1-12). We did not perform any experiments to confirm this.
We do not guarantee IHC-P for mouse. Some optimisation may be required for detection of the target protein due to low levels of endogenous expression in some samples. Please see images below for suitable positive controls.
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Tested applications
Suitable for: Flow Cyt (Intra), IHC-P, WB, ICC/IF, IPmore details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: A549, A375, HeLa, NIH/3T3, Raw264.7 cell lysates. Human fetal spleen and colon tissue lysates. Mouse spleen and bone marrow tissues lysates. IHC-P: Human cervix carcinoma, lung carcinoma and gastric carcinoma tissues, rat spleen tissue. ICC/IF: HeLa cells. Flow Cyt (intra): Jurkat cells.
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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
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 40% Glycerol, 0.05% BSA, 59% PBS -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR14639(2) -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab197896 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) |
1/250.
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IHC-P | (3) |
1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
We do not guarantee IHC-P for mouse. |
WB |
1/1000. Detects a band of approximately 12 kDa (predicted molecular weight: 12 kDa).
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ICC/IF | (2) |
1/250.
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IP |
1/40.
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Notes |
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Flow Cyt (Intra)
1/250. |
IHC-P
1/2000. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. We do not guarantee IHC-P for mouse. |
WB
1/1000. Detects a band of approximately 12 kDa (predicted molecular weight: 12 kDa). |
ICC/IF
1/250. |
IP
1/40. |
Target
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Tissue specificity
Ubiquitously expressed. -
Sequence similarities
Belongs to the S-100 family.
Contains 2 EF-hand domains. - Information by UniProt
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Database links
- Entrez Gene: 6275 Human
- Entrez Gene: 20198 Mouse
- Entrez Gene: 24615 Rat
- Omim: 114210 Human
- SwissProt: P26447 Human
- SwissProt: P07091 Mouse
- SwissProt: P05942 Rat
- Unigene: 654444 Human
see all -
Alternative names
- 18A2 antibody
- 42A antibody
- calcium Placental protein antibody
see all
Images
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lane 3 : Wild-type A549 cell lysate
Lane 4 : S100A4 knockout A549 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-S100A4 antibody [EPR14639(2)] staining at 1/1000 dilution, shown in green;loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) staining at 1/20000 dilution, shown in red. In Western blot, ab197896 was shown to bind specifically to S100A4. A band was observed at 11 kDa in wild-type HeLa and A549 cell lysates with no signal observed at this size in S100A4 knockout HeLa cell line ab265709 (knockout cell lysate ab257046) and S100A4 knockout A549 cell line ab261865 (knockout cell lysate ab261674). To generate this image, wild-type and S100A4 knockout HeLa and S100A4 knockout A549 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.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type A549 whole cell lysate
Lane 2 : S100A4 knockout A549 whole cell lysate
Lane 3 : A375 whole cell lysate
Lane 4 : HeLa whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 12 kDa
Observed band size: 12 kDaLanes 1 - 4: Merged signal (red and green). Green - ab197896 observed at 12 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab197896 was shown to recognize S100A4 in wild-type A549 cells as signal was lost at the expected MW in S100A4 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and S100A4 knockout samples were subjected to SDS-PAGE. Ab197896 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed 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 1/20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?Lanes 1 - 2: Merged signal (red and green). Green - ab197896 observed at 11 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab197896 was shown to react with S100A4 in wild-type HeLa cells in Western blot with loss of signal observed in S100A4 knockout cell line ab265709 (S100A4 knockout cell lysate ab257046). Wild-type HeLa and S100A4 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab197896 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.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 12 kDaLanes 1 - 2: Merged signal (red and green). Green - ab197896 observed at 12 kDa. Red - loading control ab8245 observed at 37 kDa.
ab197896 Recombinant Anti-S100A4 antibody [EPR14639(2)] was shown to specifically react with S100A4 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab261758 (knockout cell lysate ab257045) was used. Wild-type and S100A4 knockout samples were subjected to SDS-PAGE. ab197896 and Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-S100A4 antibody [EPR14639(2)] (ab197896)
Immunohistochemical analysis of paraffin-embedded Rat spleen tissue labeling S100A4 with ab197896 at 1/2000 dilution (0.376 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on lymphocytes of rat spleen.The section was incubated with ab197896 for 30 mins at room temperature. The immunostaining staining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins.
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).
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Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling S100A4 with ab197896 at 1/200 (10 μg/mL). Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% TritonX-100. ab150077, AlexaFluor®488 Goat anti-Rabbit at 1/1000 (2 μg/mL) was used as the secondary antibody. Cells were counterstained with ab195889, Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 (2.5 μg/mL). Nuclear counter stain was DAPI (blue).Confocal image showing positive staining in HeLa cell line.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lanes 1 & 5 : A549 whole cell lysate
Lane 2 : Human liver lysates
Lane 3 : Human lung lysates
Lane 4 : Human colon lysates
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 12 kDa
Observed band size: 12 kDa
Exposure time: 26 seconds -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-S100A4 antibody [EPR14639(2)] (ab197896)
Immunohistochemical analysis of paraffin-embedded Human gastric carcinoma tissue labeling S100A4 using ab197896 at 1/2000 dilution. Goat Anti-Rabbit IgG H&L (HRP) ab97051 was used as a secondary antibody at 1/500 dilution. Cells were counterstained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab197896 and secondary antibody only.
Note: Cytoplasm and nuclear staining on human gastric carcinoma tissue was observed.Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : A375 (human malignant melanoma) whole cell lysate
Lane 2 : Human fetal spleen lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution
Predicted band size: 12 kDa
Exposure time: 3 minutesBlocking/dilution buffer: 5% NFDM/TBST
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Intracellular Flow Cytometry analysis of Jurkat cells labelling S100A4 with ab197896 at 1/250 (red). Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. An Alexa Fluorr® 488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.
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ab197896 at 1/40 immunoprecipitating S100A4 in A549 whole cell lysate observed at 12 KDa.
Lane 1 (input): A549 whole cell lysate 10μg
Lane 2 (+): ab197896 + A549 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab197896 in A549 whole cell lysate
For western blotting, Panel A: ab197896, 1:1000; Panel B: ab124805, 1:1000 and anti-rabbit IgG (HRP), specific to the non-reduced form of IgG was used as the secondary antibody (1/1500).
Blocking/Diluting buffer and concentration: 5% NFDM/TBST.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-S100A4 antibody [EPR14639(2)] (ab197896)
Immunohistochemical analysis of paraffin-embedded Human cervix carcinoma tissue labeling S100A4 using ab197896 at 1/2000 dilution. Goat Anti-Rabbit IgG H&L (HRP) ab97051 was used ay 1/500 dilution as a secondary antibody and cells were counterstained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab197896 and secondary antibody only.
Note: Nuclear and cytoplasm staining on cervix carcinoma tissue was observed.Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-S100A4 antibody [EPR14639(2)] (ab197896)
Immunohistochemical analysis of paraffin-embedded Human lung carcinoma tissue labeling S100A4 using ab197896 at 1/2000 dilution. Goat Anti-Rabbit IgG H&L (HRP) ab97051 was used as a secondary antibody at a dilution of 1/500 and cells were counterstained with Hematoxylin.
Inset image: negative control obtained using PBS instead of ab197896 and secondary antibody only.
Note: Nuclear and weakly staining on lung carcinoma tissue was observed.Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : A549 (Human lung carcinoma epithelial cell) whole cell lysates
Lane 2 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates
Lane 3 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
Lane 4 : Mouse spleen tissue lysates
Lane 5 : Mouse bone marrow tissue lysates
Lane 6 : Mouse heart tissue lysates
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 12 kDa
Observed band size: 12 kDa
Exposure time: 44 secondsBlocking and dilution buffer: 5% NFDM/TBST.
Some optimisation may be required for detection of the target protein due to low levels of endogenous expression in some samples.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (52)
ab197896 has been referenced in 52 publications.
- Qin HY et al. Associations among S100A4, Sphingosine-1-Phosphate, and Pulmonary Function in Patients with Chronic Obstructive Pulmonary Disease. Oxid Med Cell Longev 2022:6041471 (2022). PubMed: 35165531
- Jia M et al. Inhibition of PI3K/AKT/mTOR Signalling Pathway Activates Autophagy and Suppresses Peritoneal Fibrosis in the Process of Peritoneal Dialysis. Front Physiol 13:778479 (2022). PubMed: 35309056
- Song L et al. Dexmedetomidine Protects Against Kidney Fibrosis in Diabetic Mice by Targeting miR-101-3p-Mediated EndMT. Dose Response 20:15593258221083486 (2022). PubMed: 35370507
- Li DK et al. Exosomal HMGA2 protein from EBV-positive NPC cells destroys vascular endothelial barriers and induces endothelial-to-mesenchymal transition to promote metastasis. Cancer Gene Ther 29:1439-1451 (2022). PubMed: 35388172
- Qi Y et al. Macrophage-Secreted S100A4 Supports Breast Cancer Metastasis by Remodeling the Extracellular Matrix in the Premetastatic Niche. Biomed Res Int 2022:9895504 (2022). PubMed: 35496059