Anti-IL-6 抗体 (ab6672)

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1. If you prefer to do the incubation at room temperature you should not exceed a 2 hour incubation. We do recommend an overnight incubation at 4 degrees, however, as this usually gives the strongest and most specific staining.
2. I would suggest you use ab6672 at 1/400 and ab18672 at 1/2000.
3. I believe lung cancer tissue is a good positive control for these antibodies but am not sure that the levels in normal tissue are high enough. For ab18672 we have seen good expression in lymph node, poocine epithelial tissue, and human temporal artery. For ab6672 human and mouse pancreatic cancer, head and neck tumor, and human liver sections have shown good results. Any tissue reported in the literature to have good IL6 or IL8 expression should be a good positive control if you do not have access to these tissues.
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I think ab27472 that you have selected is a great isotype control for ab6672 as it is a rabbit polyclonal IgG and ab6672 is also a rabbit polyclonal with isotype IgG.

Thank you for your enquiry regarding ab6672.

This product is not affinity purified antibody but whole antiserum. Thus the storage buffer may contain degradation product of red blood cells which can contribute to the brownish color.

Thank you for your email.

I have discussed the issue with laboratory. They have advised that the red colour is due to hemolysis of blood during processing of the serum and did not affect the function of this rabbit antiserum during testing. We normally avoid hemolysis as much as possible, but it is sometimes unavoidable, and it does not affect the function of this primary antibody.

I would reassure you that antibody is absolutely fine and would strongly recommend using it.

We unfortunately will not be able to provide this antibody in clear solution so if you do not want to use it then I would suggest choosing another antibody from catalogue. Please let me know the catalogue number and I will send you the same as replacement vial. I will also send you the details for returning the ab6672. Could you please store the antibody at -20C until I arrange a collection?

Thank you very much for your continuous cooperation. I will look forward to hearing from you soon.

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The immunogen of ab6672 is full length human IL6 which shares 40% sequence homology with rabbit IL6 while the immunogen of ab43142 shares 14.9% with rabbit IL6.

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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Optimization of antigen retrieval step in IHC-P is very important. Please try the antigen retrieval by incubating the tissue sections at different time intervals e.g. 5 minutes, 10, 15, 20, 25 and 30 minutes. This range will help to get the optimized time at which the epitopes are exposed for maximum antibody reactivity.

The secondary antibody details has not been provided. Have you used secondary antibody? Please try anti rabbit HRP/AP conjugated as required.

Please try blocking with 5% BSA or 10% normal serum for 1-2 hours.

Fixation of tissue sections in formalin or PFA for more than 24 hours actually decreases the antibody binding capacity due to cross-linking. I would recommend trying the 24hr fixed sections.

Add 0.5% Tween in antibody diluting buffer.

Please recheck the buffer pH; low or higher pH will interfere with the epitope retrieval.

I hope these suggestions will be helpful. Should you have any question please do not hesitate to contact me.

Thank you for contacting us.

Here is the recommended protocol for IHC:

The below information is only intended as a guide. The researcher should determine what protocol best meets their needs. Please follow safe laboratory procedures.

Frozen Sections

Snap-freeze fresh tissues in liquid nitrogen or isopentane pre-cooled in liquid nitrogen, embedded in OCT compound in cryomolds. Store frozen blocks at -80º C.

Cut 4-8 m m thick cryostat sections and mount on superfrost plus slides or gelatin coated slides. Store slides at - 80º C until needed.

Before staining, warm slides at room temperature for 30 minutes and fix in ice-cold acetone for 10 minutes. Air-dry for 30 minutes.

Wash in PBS



Paraffin Sections

Deparaffinize sections in xylene, 2x5min.

Hydrate with 100% ethanol, 2x3min.

Hydrate with 95% ethanol, 1min.

Rinse in ultra pure water.



Procedure for Immunoenzyme Staining

Follow procedure for pretreatment as required (for ab6672 citrate buffer, pH 6 was commonly used)

Rinse Sections in PBS for 2x2 min

Serum Blocking: incubate sections in normal serum block ? species same as secondary antibody (for example, Normal Goat Serum (NGS) if secondary antibody is goat host).
Note: since this protocol uses avidin-biotin detection system, avidin/biotin block may be needed based on tissue type. If you do, the avidin/biotin blocking should be done after normal serum block and before primary antibody incubation.

Primary Antibody: incubate sections in primary antibody at appropriate dilution in dilution buffer (PBS supplemented with serum block) for 1 hour at room temperature or overnight. Note: Do not rinse sections between serum block and primary antibody incubation.

Rinse in PBS buffer for 3x2 min.

Peroxidase Blocking: incubate sections in 1% hydrogen peroxidase in PBS for 10 minutes at room temperature.

Rinse in PBS buffer for 3x2 min.

Secondary Antibody: incubate sections in biotinylated secondary antibody (1:500, for example) in PBS buffer for 30 minutes at room temperature.

Rinse in PBS buffer for 3x2 min.

Detection: incubate sections in streptavidin-HRP (1:500, for example) in PBS buffer for 30 minutes at room temperature.

Rinse in PBS buffer for 3x2 min.

Chromagen/Substrate: incubate sections in peroxidase substrate solution. Use stable blue or brown staining, respectively.

Rinse in PBS buffer for 3x2 min.

Rinse in ultra pure water for 3x5 min.

Dehydrate through 95% ethanol for 1 minute, 100% ethanol for 2x3min.

Clear in xylene for 2x5min.

Coverslip with mounting medium.



IL6 is a secreted protein, so it should be found in the cytoplasm and/or extracellular space. http://www.uniprot.org/uniprot/P05231

You can email that reviewer by clicking on her name in the Abreview online.

https://www.abcam.com/IL-6-antibody-ab6672.html&tab=abreviews&intabreviewid=2674

https://www.abcam.com/IL-6-antibody-ab6672.html&tab=abreviews&intabreviewid=7374

I do not know if she's ever published using this antibody. I don't see her name on the list of references for this antibody: https://www.abcam.com/IL6-antibody-ab6672-references.html I'd recommend that you contact her through our Abreview system.

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

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Thank you for contacting Abcam regarding ab6672.

This antibody is a whole antiserum, thus coloration will vary a bit from lot to lot. The appearance of the antibody you received is normal, however if you have any difficulties using the antibody, we are happy to replace it per our Abpromise guarantee.

I hope this information is helpful. Please do not hesitate to contact me if you have any additional questions.