Overview

  • Product name
  • Description
    Rabbit polyclonal to Endostatin
  • Host species
    Rabbit
  • Specificity
    Detects recombinant human endostatin.
  • Tested applications
    Suitable for: ICC, ICC/IF, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Human, Non human primates
  • Immunogen

    Synthetic peptide corresponding to Human Endostatin aa 129-142.
    Sequence:

    RRLM/TESYCETWRTE


    (Peptide available as ab4983)

  • General notes

    ab3453 has been recombinant only tested in Western blot

Properties

Applications

Our Abpromise guarantee covers the use of ab3453 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 Use at an assay dependent concentration.
ICC/IF 1/200 - 1/1000.
IHC-P 1/100 - 1/500.

Target

  • Function
    COLA18A probably plays a major role in determining the retinal structure as well as in the closure of the neural tube.
    Endostatin potently inhibits endothelial cell proliferation and angiogenesis. May inhibit angiogenesis by binding to the heparan sulfate proteoglycans involved in growth factor signaling.
  • Tissue specificity
    Present in multiple organs with highest levels in liver, lung and kidney.
  • Involvement in disease
    Defects in COL18A1 are a cause of Knobloch syndrome (KNO) [MIM:267750]. KNO is an autosomal recessive disorder defined by the occurrence of high myopia, vitreoretinal degeneration with retinal detachment, macular abnormalities and occipital encephalocele.
  • Sequence similarities
    Belongs to the multiplexin collagen family.
    Contains 1 FZ (frizzled) domain.
    Contains 1 TSP N-terminal (TSPN) domain.
  • Post-translational
    modifications
    Prolines at the third position of the tripeptide repeating unit (G-X-Y) are hydroxylated in some or all of the chains.
  • Cellular localization
    Secreted > extracellular space > extracellular matrix.
  • Information by UniProt
  • Database links
  • Alternative names
    • Alpha 1 collagen type 18 (XVIII)(COL18A1) antibody
    • Alpha 1 type XVIII collagen antibody
    • Antiangiogenic agent antibody
    • COIA1_HUMAN antibody
    • COL15A1 antibody
    • Col18a1 antibody
    • Collagen alpha 1(XV) chain antibody
    • Collagen alpha 1(XVIII) chain antibody
    • Collagen alpha-1(XV) chain antibody
    • Collagen type XV proteoglycan antibody
    • Collagen type XVIII alpha 1 antibody
    • Collagen XV, alpha 1 polypeptide antibody
    • Collagen, type XV, alpha 1 antibody
    • Endostatin antibody
    • Endostatin XV antibody
    • FLJ27325 antibody
    • FLJ34914 antibody
    • FLJ38566 antibody
    • KNO antibody
    • KNO1 antibody
    • KS antibody
    • MGC74745 antibody
    • Multi functional protein MFP antibody
    • OTTHUMP00000021782 antibody
    • OTTHUMP00000115472 antibody
    • OTTHUMP00000115473 antibody
    see all

Images

  • ab3453 labelling Endostatin (green) in 293T cells (right) compared with a negative control (left) by Immunocytochemistry/Immunofluorscence. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes, blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were incubated with the primary antibody (1:200 in 3% BSA-PBS) overnight at 4 ºC. A DyLight 488-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Red (phalloidin) - F-actin, Blue (DAPI) - nuclei (blue). Images were taken at a magnification of 60x.

  • ab3453 labelling Endostatin (green) in A431 cells (right) compared with a negative control (left) by Immunocytochemistry/Immunofluorscence. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes, blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were incubated with the primary antibody (1:200 in 3% BSA-PBS) overnight at 4 ºC. A DyLight 488-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Red (phalloidin) - F-actin, Blue (DAPI) - nuclei (blue). Images were taken at a magnification of 60x.

  • ab3453 labelling Endostatin (green) in NIH-3T3 cells (right) compared with a negative control (left) by Immunocytochemistry/Immunofluorscence. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes, blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were incubated with the primary antibody (1:200 in 3% BSA-PBS) overnight at 4 ºC. A DyLight 488-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Red (phalloidin) - F-actin, Blue (DAPI) - nuclei (blue). Images were taken at a magnification of 60x.

  • ab3453 labelling Endostatin in the secretion of Human kidney tissue (right) compared with a negative control (left) by Immunohistochemistry (formalin/PFA-fixed paraffin embedded sections). To expose target proteins, antigen retrieval method was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature. Tissue sections were incuabted with the primary antibody (1:100 in 3% BSA-PBS) overnight at 4°C. A HRP-conjugated anti-rabbit IgG was used as the secondary antibody, followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

  • ab3453 labelling Endostatin in the secretion of Human colon tissue (right) compared with a negative control (left) by Immunohistochemistry (formalin/PFA-fixed paraffin embedded sections). To expose target proteins, antigen retrieval method was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature. Tissue sections were incuabted with the primary antibody (1:200 in 3% BSA-PBS) overnight at 4°C. A HRP-conjugated anti-rabbit IgG was used as the secondary antibody, followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

  • ab3453 labelling Endostatin in the secretion of Mouse kidney tissue (right) compared with a negative control (left) by Immunohistochemistry (formalin/PFA-fixed paraffin embedded sections). To expose target proteins, antigen retrieval method was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature. Tissue sections were incuabted with the primary antibody (1:100 in 3% BSA-PBS) overnight at 4°C. A HRP-conjugated anti-rabbit IgG was used as the secondary antibody, followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

References

This product has been referenced in:
  • Mohajeri A  et al. Effect of Culture Condition Variables on Human Endostatin Gene Expression in Escherichia coli Using Response Surface Methodology. Jundishapur J Microbiol 9:e34091 (2016). Read more (PubMed: 27800134) »
See 1 Publication for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Answer

Thank you for your reply.


Yes, my apologies, the µ symbol did not paste correctly from the protocol. 100 ug were resolved on 10% SDS gels under reducing conditions.


The recommended working dilution of the antibody is 2ug/ml. You should try incubating with the primary at 2ug/ml overnight at 4C and perhaps increasing the amount of protein loaded if you can.


If the protocol suggestions do not improve your results, we will replace or refund the product for you.


I look forward to hearing how today's WB worked and I'm sorry you're not feeling well! We'll get this sorted out for you.

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Question
Answer

Thank you for contacting us. I am sorry to hear that this antibody is not providing satisfactory results.

The lab had tested the below protocol:

Tumours injected 2 days earlier with angiostatin or empty expression plasmids were excised, minced with scissors, and homogenized in protein lysate buffer [50 mmol /LTris, pH 7.4, 100 mol/L EDTA, 0.25 mol/L sucrose, 1% sodium dodecyl sulfate (SDS), 1% NP40, 1 g/mL leupeptin, 1 g/mL pepstatin A, and 100 mol/L phenylmethylsulfonyl fluoride ] at 48C using a motordriven Virtus homogenizer (Virtus, Gardiner, NY). Tumour lysates from each group of mice were pooled, and protein content was determined. Tissue or cell debris was removed by centrifugation at 10,000g for 10 minutes at 48C. Protein samples (100 g) were resolved on 10% polyacrylamide SDS gels under reducing conditions, and electrophoretically transferred to nitrocellulose Hybond C extra membranes (Amersham Life Science, Buckingham, England, UK). After blocking the membranes with 5% bovine serum albumin in TTBS (Tween 20/Tris - buffered saline 20 mmol /L Tris, 137 mmol/L NaCl, pH 7.6, containing 0.1% Tween-20), blots were incubated with endostatin antibody at 2 ug/ml overnight at 4C. They were then incubated with horseradish peroxidase–conjugated secondary antibody, and developed by enhanced chemiluminescence (Amersham International, Buckingham, England, UK) and exposure to x-ray film. Band density was quantified using Scion.


You may want to try changing your blocking buffer to 5% BSA instead of milk.


Also, what gel percentage were you using? It may bebest to try 10% gel if you haven't already since the protein of interest is so small.


Did you also use a loading control, like beta actin, or check the transfer with Ponceau?


In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

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