Overview

  • Product name
  • Description
    Rabbit polyclonal to IL-8
  • Host species
    Rabbit
  • Specificity
    Please be aware that this product has low homology with the mouse and rat sequence of IL8 (Rat, 46%; Mouse 45%, UniProt blast) and we therefore cannot guarantee reactivity in these species.
  • Tested applications
    Suitable for: IHC-P, WB, IHC-Fr, IP, ELISA, ICC/IFmore details
  • Species reactivity
    Reacts with: Human, Cynomolgus monkey, Rhesus monkey
  • Immunogen

    Recombinant full length protein corresponding to IL-8.

  • Positive control

Properties

Applications

Our Abpromise guarantee covers the use of ab7747 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
IHC-P Use at an assay dependent concentration. PubMed: 17208399
WB 1/10. Detects a band of approximately 6-8 kDa (predicted molecular weight: 11.1 kDa). IL-8 appears to exist as a dimer in solution.
IHC-Fr Use at an assay dependent concentration. PubMed: 25120588
IP 1/10.
ELISA Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration. PubMed: 26270987

Target

  • Function
    IL-8 is a chemotactic factor that attracts neutrophils, basophils, and T-cells, but not monocytes. It is also involved in neutrophil activation. It is released from several cell types in response to an inflammatory stimulus. IL-8(6-77) has a 5-10-fold higher activity on neutrophil activation, IL-8(5-77) has increased activity on neutrophil activation and IL-8(7-77) has a higher affinity to receptors CXCR1 and CXCR2 as compared to IL-8(1-77), respectively.
  • Sequence similarities
    Belongs to the intercrine alpha (chemokine CxC) family.
  • Post-translational
    modifications
    Several N-terminal processed forms are produced by proteolytic cleavage after secretion from at least peripheral blood monocytes, leukcocytes and endothelial cells. In general, IL-8(1-77) is referred to as interleukin-8. IL-8(6-77) is the most promiment form.
  • Cellular localization
    Secreted.
  • Information by UniProt
  • Database links
  • Alternative names
    • (Ala-IL-8)77 antibody
    • (Ser-IL-8)72 antibody
    • 9E3 antibody
    • Beta thromboglobulin like protein antibody
    • C-X-C motif chemokine 8 antibody
    • CEF-4 antibody
    • chemokine, CXC motif, ligand 8 antibody
    • CXCL8 antibody
    • Emoctakin antibody
    • GCP-1 antibody
    • GCP/IL-8 protein I antibody
    • GCP/IL-8 protein II antibody
    • GCP/IL-8 protein III antibody
    • GCP/IL-8 protein IV antibody
    • GCP/IL-8 protein V antibody
    • GCP/IL-8 protein VI antibody
    • GCP1 antibody
    • Granulocyte chemotactic protein 1 antibody
    • IL-8 antibody
    • IL-8(1-77) antibody
    • IL-8(9-77) antibody
    • IL8 antibody
    • IL8/NAP1 form I antibody
    • IL8/NAP1 form II antibody
    • IL8/NAP1 form III antibody
    • IL8/NAP1 form IV antibody
    • IL8/NAP1 form V antibody
    • IL8/NAP1 form VI antibody
    • IL8_HUMAN antibody
    • Inteleukin 8 antibody
    • LECT antibody
    • LUCT antibody
    • Lymphocyte-derived neutrophil-activating factor antibody
    • LYNAP antibody
    • MDNCF antibody
    • MDNCF-b antibody
    • MDNCF-c antibody
    • MONAP antibody
    • Monocyte-derived neutrophil chemotactic factor antibody
    • Monocyte-derived neutrophil-activating peptide antibody
    • NAF antibody
    • NAP-1 antibody
    • NAP1 antibody
    • Neutrophil activating peptide 1 antibody
    • Neutrophil activating protein 1 antibody
    • Neutrophil-activating factor antibody
    • Neutrophil-activating protein 1 antibody
    • Protein 3 10C antibody
    • Protein 3-10C antibody
    • SCYB8 antibody
    • Small inducible cytokine subfamily B member 8 antibody
    • T cell chemotactic factor antibody
    • T-cell chemotactic factor antibody
    • TSG1 antibody
    see all

Images

  • ab7747 staining IL8 in SVGA astrocytes transfected with a plasmid encoding Vpr by ICC/IF (Immunocytochemistry/immunofluorescence). The cells were stained for nucleus (blue), IL8 (green) and GFAP (red). Cells were fixed with ice-cold methanol/acetone (1:1) for 20 minutes at 20°C, washed 3x with PBS and permeabilized with 0.1% Triton X-100 in PBS and blocked with 1% BSA for 30 minutes at RT. Samples were incubated with ab7747 overnight in a humidified chamber. The cells were washed 5x with 0.1% Triton X-100 in PBS and incubated for 1h in secondary antibodies conjugated with Alexa Fluor 488 (Anti-Rabbit, 1:1000) at room temperature in the dark. All the antibodies were diluted in PBS containing 1% BSA.

References

This product has been referenced in:
See all 35 Publications for this product

Customer reviews and Q&As

1-10 of 11 Q&A

Answer

Vielen Dank für Ihren Anruf und für Ihr Interesse an unseren Produkten.

Unseres Wissens nach wurde ab7747 bisher noch nicht in Durchflusszytometrie getestet. Falls Sie dies selbst testen möchten, kann ich Ihnen zur Zeit ein spezielles Angebot über einen 100%igen Abreview-Rabatt anbieten. Bei diesem Angebot bekommen Sie einen Rabatt für eine zukünftige Bestellung, wenn Sie uns ein Abreview mit dem Testresultat zusenden. Der Rabatt würde den ganzen Wert von ab7747 abdecken, und gegen eine erneute Bestellung weitere Produkte von uns verrechnet werden.

Um von diesem Angebot profitieren zu können, folgen Sie bitte diesen Schritten:

1.) Bestätigen Sie mir bitte, dass Sie ab7747 in Durchflusszytometrie testen möchten.

2.) Bitte bestellen Sie erst nach Erhalt des Rabattcodes.

3.) Bestellen Sie den Antikörper wie üblich per Telefon, Email oder Fax

4.) Testen Sie den Antikörper in Durchflusszytometrie.

5.) Teilen Sie uns das Ergebnis Ihres Tests durch ein Abreview mit und notieren Sie den Discount Code in dem Feld "additional notes"
Unter der folgenden URL können Sie mehr über unser Abreview System erfahren:
https://www.abcam.com/abreviews

6.) Der Rabattcode ist nach dem Abschicken des Abreviews aktiv, und Sie könnenweitere Produkte zu dem gleichen Preis wie ab7747 bei uns bestellen (halten Sie bei der Bestellung bitte den Rabattcode und die Bestellnummer von ab7747 bereit). Bitte beachten Sie, dass der Rabattcode innerhalb von 4 Monaten nach Ausstellung eingelöst werden muss.

Der Rabattcode wird gültig unabhängig davon, ob Ihr Ergebnis positiv oder negativ ist.

Die Bedingungen zu unserem 100% Abreview Rabatt können Sie unter dem folgenden Link nachlesen:
https://www.abcam.com/collaborationdiscount

Ich hoffe, diese Informationen helfen Ihnen weiter. Falls Sie einen Rabattcode erhalten möchten oder weitere Fragen haben, zögern Sie bitte nicht, sich wieder an mich zu wenden.

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Answer

Thank you for contacting us. Because we carry over 70,000 products, it isn't feasible for us to keep small sample sizes of our products.

We are happy to reassure our customers that all of our products are covered by our Abpromise, which guarantees that the product will work in the applications and species specified on the datasheet, or we will offer a replacement, credit, or refund within 6 months of purchase.

If the product is to be used in an untested species or application, you may be eligible for our testing discount program if the antibody has not yet been purchased. Please contact our Scientific Support team by replying to this email prior to purchase for more information.

Otherwise, we like to encourage all of our customers to submit an Abreview via the online product datasheet. We always appreciate customer feedback, whether positive or negative, and we make all product information available to researchers. Plus, each Abreview earns Abpoints that can be used for discounts on future purchases or rewards such as Amazon.com gift certificates.

To find out more about our Abreview system, please see the following link:

https://www.abcam.com/abreviews

I hope this information is helpful. Please do not hesitate to contact us again with any other questions.

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Answer

Thank you for your reply.

In theory it is possible to perform IHC-P with a directly conjugated antibody. This will also mean that the amplification that is achieve by adding the secondary is lost.

So my advise would be to only do it for abundant antigens.

ab7747 is not conjugated to HRP and therefore will need a secondary antibody that is conjugated.

I can recommend to use one of these:

ab98485

https://www.abcam.com/index.html?datasheet=98485 (or use the following: https://www.abcam.com/index.html?datasheet=98485).

ab97085

https://www.abcam.com/index.html?datasheet=97085 (or use the following: https://www.abcam.com/index.html?datasheet=97085).

We also offer EasyLink kits that can be used to conjugate this antibody directly to HRP (ab102892).

https://www.abcam.com/index.html?datasheet=102892 (or use the following: https://www.abcam.com/index.html?datasheet=102892).

It would not be 100% optimal in this case since the antibody is only purified as IgG fraction.

I hope this information is helpful and wish you a good weekend.

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Answer

Thank you for your inquiry.

I am happy to confirm that the recommended protocol in IHC is the following:

Antigen retrieval: 5 min proteolytic enzyme (competitor), 20C

Antibody incubation: 120 min, 20C 1:500

(http://www.ncbi.nlm.nih.gov/pubmed/17208399?dopt=Abstract)

I hope this information is helpful and wish you good luck with your experiments.

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Question
Answer

Thank you for your call today and for letting us know about the trouble with ab7747.

As we discussed, I'm sending a free of charge vial of ab18672 on the order *** which should arrive tomorrow.

Please keep me updated about the results using this new antibody. I look forward to hearing from you, and let me know if you have any questions or if there is anything else that we can do for you.

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Question
Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number To check the status of the order please contact our Customer Service team and reference this number. Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know. I wish you the best of luck with your research.

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Question

Sorry about that - here it is. Primary antibody: ab7747 Secondary antibody: Biotinylated goat anti-rabbit IgG [a competitor] Chromogen: AEC Procedure: 1. Place slides on hot plate, 65 C for 30 minutes, deparaffinize in xylenes 4 min x 2, hydrate through alcohol –100% 2 min x 2, and 95% alcohol 2min x 2, 80% alcohol 2min x 1, then wash with PBS 3 min x2. 2. Perform antigen retrieval: Dako target retrieval buffer. Place slides in coplin jar filled with 1X antigen retrieval buff (DAKO). Pre-heat buffer for 20 minutes in steamer. Add tissue for 20 min, then leave in steamer to cool down for 30 min. 3. Draw a hydrophobic circle around the tissue with a pap pen. Add a drop of PBS and leave in humidity chamber. 4. Block endogenouse peroxides with 3% H2O2 in PBS for 12 min. 5. Wash with PBS 3min x 3. 6. Incubate with protein clocking solution (5% normal horse serum + 1% normal goat serum in PBS) 20 min. 7. Remove excess protein block soln, and add primary antibody in protein block solution (see antibody dilution schedule) and incubate overnight at 4 C. 8. Day 2, wash with PBS 3min x 3. 9. Add protein block solution and incubate for 10 min. 10. Remove excess protein block soln, add secondary antibody: biotinylated goat anti-rabbit IgG (Biocare Medical) 30 min, 11. Wash with PBS 3min x3 12. Strepavidin conjugated with horseradish peroxidase (Dako) in PBS (1:300) for 30 min. 13. Wash with PBS 3min x 3, followed by a brief rinse in dH2O with brij (1 drop of brij in 50 ml of PBS) 14. Add chromogen (AEC) and incubate for 2-5 min at RT. Check the reaction microscopically to determine the end point. 15. Wash with dH2O and dry on heat 65 C for 10-20 min, then mount with universal or Permont (Fisher) mounting solution. Leave slides overnight for a complete dry.

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Answer

Thank you for your protocol, I have looked at it in details and I would like to suggest trying the following changes to see if you start having a cytoplasmic staining rather than a nuclear staining. -check you are not overfixing (or underfixing) the tissue, different fixation times could be tested to see if this may be a problem -try a different antigen retrieval method, such as enzymatic. This way if the citrate buffer antigen retrieval method is damaging the epitope (and other proteins) you may see a difference with a different antigen retrieval step -check with a no primary control that the staining is not due to the secondary or other further steps in the staining protocol -try blocking peroxidases with only 0.3% 10-20min, too much peroxidase can damage the tissue. -remove the horse serum and use only 10% normal goat serum to block the tissue (just in case the serum is giving the problem) I think in general your protocol is very good and hope that the above suggestions will help. If you want to be certain of the staining I would recommend to run another IL8 antibody along the sister sections to see if it gives you the same staining pattern. As ab7747 has not been tested in IHC it is hard to be sure the problem is not that the antibody does not work in IHC.

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Answer

That is great to hear. Please contact us again with any more questions or comments.

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Answer

The originator of ab7747 has suggested trying recombinant IL-8 to test your system or using activated macrophages. Since you don't have the original vial, do you have the Abcam order number or the purchase order number? Do you happen to know the lot number of the previous batch that you received which worked for you?

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Question

ANTIBODY CODE ab7747 BATCH NUMBER not sure ORDER NUMBER not sure DESCRIPTION OF THE PROBLEM No signal detected by 2 users. SAMPLE Cell lysates collected from human prostate cancer cell lines PRIMARY ANTIBODY Abcam Ab7747: dilution of 100 microlitres in 15ml TBST + 5% marvel, incubation overnight at 4 degrees. Washes: 1x15 mins and 3x5 mins in TBST. These conditions have worked before. SECONDARY ANTIBODY anti-rabbit, 1:2000 dilution in TBST +5% marvel, incubated for 1 hour at room temp. Washes as before These conditions have worked before. DETECTION METHOD Have tried ECL, ECLPlus and Supersignal. None of these give a band at the correct size. POSITIVE AND NEGATIVE CONTROLS USED Samples that gave a positive result with a different batch of antibody. Cells treated with siRNA against IL8 as negative control ANTIBODY STORAGE CONDITIONS Aliquotted at -20 celcius SAMPLE PREPARATION RIPA with PI tablet + sodium orthovanadate ELB with PI tablet Samples mixed with loading buffer + mercaptoethanol & heated to 95 degrees for 5 minutes. AMOUNT OF PROTEIN LOADED 40 micrograms ELECTROPHORESIS/GEL CONDITIONS 20% reducing gel These conditions have worked before. TRANSFER AND BLOCKING CONDITIONS Transfer for 2 hours in transfer buffer containing 40% methanol. Membrane checked using Ponceau-s. Blocked in TBST +5% marvel for 2 hours. These conditions have worked before. HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes ADDITIONAL NOTES I have tried the western 3 times and another user has tried twice. I have tried samples that gave a positive result with a previous batch of antibody. Different sample preparation buffers give no different result. e see a non-specific band arounf 40kDa (have seen this before) but no IL8 band.

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Answer

Thank you for your patience. I am working with the originator of ab7747 to help determine what may be going on with this particular antibody. I would like to confirm, with this batch of ab7747 you never saw an Il-8 band and the only band you saw was at approximately 40 kDa, correct? Did you ever try other positive cells as activated macrophages? Can you give me the lot number that you received - it will be on the vial. Many thanks.

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1-10 of 11 Q&A

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