HRP Anti-Fluorescein antibody (ab6656)

Reviews (2)Q&A (7)References (6)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - HRP Anti-Fluorescein antibody (ab6656)
  • Immunohistochemistry (Frozen sections) - HRP Anti-Fluorescein antibody (ab6656)

Key features and details

  • HRP Goat polyclonal to Fluorescein
  • Suitable for: ELISA, IHC-P, IHC-Fr, WB
  • Reacts with: Species independent
  • Conjugation: HRP
  • Isotype: IgG

Overview

  • Product name

    HRP Anti-Fluorescein antibody
    See all Fluorescein primary antibodies

  • Description

    HRP Goat polyclonal to Fluorescein

  • Host species

    Goat

  • Conjugation

    HRP

  • Specificity

    This antibody reacts with FITC-conjugated proteins.

  • Tested applications

    Suitable for: ELISA, IHC-P, IHC-Fr, WBmore details

  • Species reactivity

    Reacts with: Species independent

  • Immunogen

    Fluorescein conjugated to Goat IgG

  • General notes

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Store at +4°C.

  • Storage buffer

    Preservative: 0.01% Gentamicin sulphate
    Constituents: 0.42% Potassium phosphate, 0.87% Sodium chloride, 1% BSA
  • Concentration information loading...

  • Purity

    Affinity purified

  • Purification notes

    This product was prepared from monospecific antiserum by immunoaffinity chromatography using Fluorescein IgG coupled to agarose beads followed by solid phase adsorption(s) to remove any unwanted reactivities.

  • Clonality

    Polyclonal

  • Isotype

    IgG

  • Research areas

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab6656 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
1/200000.
IHC-P (1)
1/500 - 1/2500.
IHC-Fr (1)
Use at an assay dependent concentration.
WB
1/1000 - 1/5000.
Notes
ELISA
1/200000.
IHC-P
1/500 - 1/2500.
IHC-Fr
Use at an assay dependent concentration.
WB
1/1000 - 1/5000.

Target

  • Relevance

    Fluorescein is a fluorophore commonly used to label proteins - protein-fluorescein conjugates are not usually susceptible to precipitation. In addition to its relatively high absorptivity, excellent fluorescence quantum yield and good water solubility, fluorescein has an excitation maximum of 494 nm that closely matches the 488 nm spectral line of the argon-ion laser, making it an important fluorophore for confocal laser-scanning microscopy and flow cytometry applications. Its fluorescence is pH sensitive and is significantly reduced below pH 7. Fluorescein emits most strongly between 500 and 550 nm, but it has a relatively broad emission spectrum reaching to over 600 nm. Several derivatives of fluorescein are commonly used, including FITC (fluorescein isothiocyanate), carboxylates and succinimidyl esters.

  • Alternative names

    • FI antibody
    • FITC antibody
    • fluorescein isothyocianate antibody
    • fluorescyl antibody

Images

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - HRP Anti-Fluorescein antibody (ab6656)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - HRP Anti-Fluorescein antibody (ab6656)This image is courtesy of an anonymous Abreview

    Immunohistochemical analysis of Mouse thymus in which the medullary epithellai cells are labelled with fluorescein-conjugated UEA1 lectin with ab6656. Tissue was fixed with paraformaldehyde and blocked with 1% BSA for 40 minutes at 20°C. Samples were incubated with primary antibody (1/1000) for 1 hour at 20°C.

    See Abreview

  • Immunohistochemistry (Frozen sections) - HRP Anti-Fluorescein antibody (ab6656)
    Immunohistochemistry (Frozen sections) - HRP Anti-Fluorescein antibody (ab6656)This image is courtesy of an anonymous Abreview

    ab6656 staining GFP in Mouse mammary carcinoma metastasis tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Perfusion fixed tissue with 4% paraformaldehyde, blocked with 0.5% Perkin-Elmer TNB blocking reagent for 30 minutes at 25°C  and permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/500) for 30 minutes at 25°C

    Micrograph demonstrates use of ab6662 for detection of GFP-positive tumor cells (LEFT) and an adjacent section after amplification with ab6656 (RIGHT). Images were taken with identical settings on a Leica confocal microscope.

    See Abreview

References (6)

Publishing research using ab6656? Please let us know so that we can cite the reference in this datasheet.

ab6656 has been referenced in 6 publications.

  • Healing E  et al. A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway. Nucleic Acids Res 47:e61 (2019). PubMed: 30869144
  • de Lucas M  et al. Transcriptional Regulation of Arabidopsis Polycomb Repressive Complex 2 Coordinates Cell-Type Proliferation and Differentiation. Plant Cell 28:2616-2631 (2016). In situ hybridization ; Arabidopsis thaliana . PubMed: 27650334
  • Lin WR  et al. The Influence of Bone Marrow-Secreted IL-10 in a Mouse Model of Cerulein-Induced Pancreatic Fibrosis. Biomed Res Int 2016:4601532 (2016). PubMed: 27314021
  • Xiang D  et al. Superior Performance of Aptamer in Tumor Penetration over Antibody: Implication of Aptamer-Based Theranostics in Solid Tumors. Theranostics 5:1083-97 (2015). IHC . PubMed: 26199647
  • Shigdar S  et al. The use of sensitive chemical antibodies for diagnosis: detection of low levels of EpCAM in breast cancer. PLoS One 8:e57613 (2013). IHC-P ; Human . PubMed: 23460885
  • Kett LR  et al. LRRK2 Parkinson disease mutations enhance its microtubule association. Hum Mol Genet 21:890-9 (2012). PubMed: 22080837

Customer reviews and Q&As

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1-9 of 9 Abreviews or Q&A

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Fluorescein antibody (HRP)

 Good
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (thymus)
Antigen retrieval step
None
Permeabilization
Yes - Tween20
Specification
thymus
Blocking step
BSA as blocking agent for 40 minute(s) · Concentration: 1% · Temperature: 20°C
Fixative
Paraformaldehyde
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Abcam user community

Verified customer

Submitted Feb 29 2016

Question

I am wondering whether this antibody (Anti-Fluorescein antibody HRP ab6656) can bind to other fluorescein derivative such as 6'-FAM. Any data upon which part of FITC is recognized by this antibody?

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Answer

Thank you for your inquiry and support of our products. The antibody is only tested and guaranteed against derivatives of FITC (Fluorescein isothiocyanate) but it may react with other forms as well. It has not been tested against 6’-FAM. The antibody is polyclonal and is expected to react with different portions of the molecule, but the specific binding sites are not characterized.

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Question

We need an anti FITC –HRP conjugated antibody to to convert FITC fluorescence to a chromogenic substrate for light microscopy (e.g., DAB) in paraffin coupes of murine mammary glands .


Both ab 6656 and ab 19492 works on IHC-par among datasheet but which antibody would you advice us to use for our application?


Is there a difference in sensitivity because ab 19492 refers to a paper in 2011 who uses an additional tyramide signal amplification system and ab 6656 refers to a very short abreview without image saying it was difficult to find a good dilution but finally it worked.

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Answer

Thank you for contacting us.

Without a direct comparison, it is a little difficult to choose between the two. The ab6656 datasheet also has a review of IHC with TSA amplification on frozen tissue, after applying the antibody at a dilution of 1/500.

Neither antibody has received any complaints in the past year but I recommend ab6656 based on the dilutions recommended for the other applications, for instance 1/1000 - 1/5000 for western blotting versus 1/20 - 1/200 for ab19492. This suggests that ab6656 has higher affinity and/or HRP activity, but we cannot be sure without direct comparison using the same samples and supplementary reagents.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Question

Goodevening, my antibody ordered over to you (order #971173) just arrived (thank you for your efficiency by the way) and I was looking for the datasheet on your website since I was searching for the recommended concentration to use it in an IP and WB assay but I can't find this information: am I missing it or it's not there? Can you help me in any way? The antibody is ab6656 anti fluorescein. Thank you very much  

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Answer

Thank you for your enquiry. The concentration of this antibody is 1.000 mg/ml, however the working dilution/concentration needs to be determined since the signal depends on several factors (such as expression levels of the target, sample preparation etc.). I would suggest that you should carry out a preliminary experiment with different dilutions of this antibody (i.e.1/100-1/1000) and then adjust the dilution accordingly from the results obtained. I hope this helps and if I can assist further, please do not hesitate to contact me.      

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Question

Will this antibody react with FITC too?

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Answer

Thank you for your phone call today and for your patience while I was in contact with the lab. The lab confirmed that ab6656 has been tested on FITC-conjugated proteins, and so the antibody is appropriate for your studies. I hope this information is helpful, but please let me know if you have any questions.  

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Immunohistochemistry (Frozen sections) abreview for Anti-Fluorescein antibody (HRP)

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (mouse mammary carcinoma metastasis)
Specification
mouse mammary carcinoma metastasis
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton X-100
Blocking step
Perkin-Elmer TNB Blocking Reagent as blocking agent for 30 minute(s) · Concentration: 0.5% · Temperature: 25°C
Read More

Abcam user community

Verified customer

Submitted May 01 2009

Question

Hi. My lab works with a class of cell-permeable, sequence-specific, DNA-binding, synthetic small molecules. One of our active compounds has a fluorescein moiety on it. Would it be possible to perform ChIP or ChIP on chip with a fluorescein antibody? Thanks.

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Answer

Thank you for your enquiry. Yes, I would consider it highly likely that you will be able to use an anti-fluorescein antibody to immunoprecipitate cross linked chromatin associated with your small molecule. You may also wish to consider NChIP as this has been employed using proteins that directly bind DNA. Furthermore if the context of chromatin is not important you may wish to adopt a DNA IP approach, incubating your small molecule with purified DNA and then performing a modified IP approach. For further details of these approaches please see our protocols web page at: http://ops.abcam.com/index.html?pageconfig=popular_protocols Your assay may require some optimisation given a potential obscuring of the fluorescein epitope under cross linked conditions, but in theory there is no reason why this should not work. Unfortunately we do not have a fluorscein antibody that has been tested using ChIP. In this case I would recommend an antibody that has been characterized and demonstrated to work in as many applications as possible, in particular IP. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

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Question

I just want to confirm that it is the correct procedure to use the HRP chromagen kit (AEC)after using the HRP conjugated fluorescein antibody?

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Answer

Thank you for your enquiry. Yes, the chromogen AEC kit is used after using the HRP conjugated fluorescein antibody in immunohistochemical techniques. If you are doing ELISA then a TMB kit is better.

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Question

I've come across the product: anti-fluorescein conjugated to HRP (product code ab6656-100). Please would it be possible to know what is the specific activity (in units / mg product) of the conjugate? I've looked in the product datasheet but this important piece of information is missing.

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Answer

Sorry, we do not have any data on the specific activity of the HRP.

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