Anti-MUC5B 抗体 [19.4E] (ab77995)

The protocol for deglycosylation is in the paper by Cao et al.
"Mucins (MUC1 and MUC3) of gastrointestinal and breast epithelia reveal different and heterogeneous tumor-associated aberrations in glycosylation"
Cao, Y, et al (1997) J Histochem Cytochem, 45:1547-57 (1997)

I have received information from the lab regarding our anti-MUC5B antibody. The immunogen is N terminus synthetic peptide of the MUC5AC tandem repeat, no glycosylation. Exact peptide sequence is proprietary and thus unavailable. Unfortunately the peptide itself is also not available for sale.

Thank you for contacting us.

Please check the following publication for deglysosylation protocol;

http://jhc.sagepub.com/content/45/11/1547.full.pdf+html

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

Thank you for contacting us.

I have been searching the antibodies as per your requirements. We have different anti Mucin 5AC antibodies however I am not sure which would better suits you because is not very clear from the immunogen.

Regarding the anti MUC 5B we have ab105460 which detects the non glycosylated domain of MUC5B. Please check he datasheet.

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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Thank you for your inquiry.


There is a protocol for deglycosylation at this website (http://www.glycotech.com/protocols/Proto3.html) and a discussion of these related to Mucin can be found in Cao, Y. et al. (1997) (http://www.ncbi.nlm.nih.gov/pubmed/9358856).


You may also opt to use a deglycosylation kit instead. I believe that Prozyme and Calbiochem carry some.


I hope this information helps. Please contact us with any other questions.

Thank you for your patience. I have received some feedback from the originator of the product. It semms that for Western Blot (and Immunohistochemistry), there is a need to perform deglycosylation in order to expose the epitope. There is a protocol for deglycosylation at this website (http://www.glycotech.com/protocols/Proto3.html) and a discussion of these related to Mucin can be found in Cao, Y. et al. (1997) (http://www.ncbi.nlm.nih.gov/pubmed/9358856). You may also opt to use a Deglycosylation kit instead. Please see attached PDF file for supplier information. Here some tips/suggestion based on the protocol you provided. 1) The antibody should be diluted using simple buffer like PBS or TBS only. 2) Was the secondary antibody used a sheep anti-human IgG antibody? Since the primary antibody was made from mouse, the secondary should be an anti-mouse IgG secondary.   I hope the suggestions above will be helpful. If not, I will be more than happy to offer you a free replacement with a different lot, or entirely different target primary antibody of your choice, if the product was purchased within the last 6 months. I look forward to hearing from you soon in order to resolve this matter. Have a nice day!  

Thank you for contacting us, we have determined that there is no cross reactivity with MUC5A or MUC5C.  I hope this helps, please let me know if you have any additional questions or concerns. 

Thank you for your call today and for your enquiry about ab77995. The dilution optimization that you mentioned on the phone sounds like a good idea to start with. I discussed the case with my colleagues, and it does sound like something in the sample might be interfering with the assay. What is used as the lavage fluid? Is it only sterile saline or PBS? Is the lavage fluid further processed in any way before being used in the assay? Are the patients healthy? You can run some tests in order to see what part of the assay is being interfered with, and then further optimization may be possible. Run the assay in one or two wells without the primary antibody, to see if there is something in the sample that non-specifically binds to the secondary antibody (i.e. Fc recepetors on white blood cells or endogenous IgG). Also have a well without the HRP-conjugated secondary antibody, to determine if there might be endogenous peroxidases in the sample. You could also run the assay using the pre-lavage fluid to make sure nothing in that solution is interfering. Once the problem is more clear, it will be easier to fix the problem. I look forward to hearing from you. Please let me know if you have any questions or if there is anything else that you need.

Thank you for contacting Abcam Technical Team and for taking the time to provide some useful details of the experiments. It is important to mention and to emphasize that the human mature MUC5B protein is heavily glycosylated and it is very likely that this is the case with the murine MUC5B as well during processing. Please advise your customer to take a look at this site for further information regarding post-translational modification: http://www.uniprot.org/uniprot/Q9HC84 http://www.uniprot.org/uniprot/Q8K1G6 If you wish me to take this enquiry further, I would appreciate if you could specify sizes of the non-specific bands and provide an image. I hope this will be useful for you. Should you still have any problem with this antibody after following these suggestions, then please do not hesitate to contact our Technical Department again.