Overview

  • Product name

    Anti-MUC5B antibody [19.4E]
    See all MUC5B primary antibodies
  • Description

    Mouse monoclonal [19.4E] to MUC5B
  • Host species

    Mouse
  • Specificity

    Does not react with human Mucin 5A or 5C.

  • Tested applications

    Suitable for: IHC-Fr, WB, ELISA, IHC-Pmore details
  • Species reactivity

    Reacts with: Mouse, Human, Monkey
  • Immunogen

    Synthetic peptide corresponding to Human MUC5B (N terminal). Un-glycosylated, serine and threonine free synthetic peptide from the N terminal region of human MUC5B.

  • General notes

    This product was changed from ascites to tissue culture supernatant on 31st May 2019. Please note that the dilutions may need to be adjusted accordingly. If you have any questions, please do not hesitate to contact our scientific support team.

Properties

Applications

Our Abpromise guarantee covers the use of ab77995 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-Fr Use at an assay dependent concentration.
WB Use at an assay dependent concentration. Predicted molecular weight: 590 kDa.
ELISA Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.

Deglycosylating the tissue is critical to expose epitope.

Target

  • Function

    Gel-forming mucin that is thought to contribute to the lubricating and viscoelastic properties of whole saliva and cervical mucus.
  • Tissue specificity

    Expressed on surface airway epithelia. Expressed mainly in mucous cells of submucosal glands of airway tissues. Highly expressed in the sublingual gland. Also found in submaxillary glands, endocervix, gall bladder, and pancreas.
  • Sequence similarities

    Contains 1 CTCK (C-terminal cystine knot-like) domain.
    Contains 3 HAT repeats.
    Contains 3 TIL (trypsin inhibitory-like) domains.
    Contains 3 VWFC domains.
    Contains 4 VWFD domains.
  • Domain

    The cysteine residues in the Cys-rich subdomain repeats are not involved in disulfide bonding.
  • Post-translational
    modifications

    Highly glycosylated. C-, N- and O-glycosylated. C-mannosylated in the Cys-rich subdomains probably on the first Trp residue of the WXXW motif. Highly O-glycosylated in the Ser/Thr-rich tandem repeat (TR) region. The repeat region is about 59% O-glycosylated with a high abundance of NeuAc(2)Hex(1)HexNac1-ol.
  • Cellular localization

    Secreted.
  • Information by UniProt
  • Database links

  • Alternative names

    • 2300002I04Rik antibody
    • A130042M24 antibody
    • AV085033 antibody
    • Cervical mucin antibody
    • Cervical mucin MUC5B antibody
    • High molecular weight salivary mucin MG1 antibody
    • MG1 antibody
    • MUC-5B antibody
    • MUC5 antibody
    • MUC5B antibody
    • MUC5B_HUMAN antibody
    • MUC9 antibody
    • Mucin 5 subtype B, tracheobronchial antibody
    • Mucin 5B antibody
    • Mucin 5B oligomeric mucus/gel forming antibody
    • Mucin-5 subtype B antibody
    • Mucin-5B antibody
    • Sublingual gland mucin antibody
    • tracheobronchial antibody
    see all

Images

  • ab77995 staining MUC5B in murine lung tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue was fixed with formaldehyde and a heat mediated antigen retrieval step was performed using citrate buffer pH 6.0. Samples were then incubated with the primary antibody at 2µg/ml for 16 hours at 4°C. MUC5B was revealed in black and nucleus are counterstain in blue (Fastblue).

    This image was generated using the ascites version of the product.

  • ab77995 staining MUC5B in murine asthma lung tissue by Immunohistochemistry (Frozen sections). Tissue was fixed with formaldehyde, followed by incubation with the primary antibody at 4µg/ml for 16 hours at 4°C. ab77995 was coupled to biotin and Streptavidin 488 (1/250) was used to amplify the signal. Counterstained with DAPI (blue).

    This image was generated using the ascites version of the product.

References

This product has been referenced in:

  • Galoian K  et al. Toll like receptors TLR1/2, TLR6 and MUC5B as binding interaction partners with cytostatic proline rich polypeptide 1 in human chondrosarcoma. Int J Oncol 52:139-154 (2018). Read more (PubMed: 29138803) »
  • Bücker R  et al. Establishment of a Mucin Secreting Cell Line Cx-03 from an Uterine Carcino Sarcoma. Pharm Res 36:7 (2018). Read more (PubMed: 30411161) »
See all 6 Publications for this product

Customer reviews and Q&As

1-10 of 11 Abreviews or Q&A

Question
Answer


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)

Read More

Answer

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.

Read More

Question
Answer

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.

Read More

Question
Answer

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.

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

Read More

Answer

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.

Read More

Question

Please help us with the ab77995 Anti-MUC5B antibody [19.4E]. Lately, we have experienced similar problem of the same nature with ab36362 Anti-Lysozyme antibody [BGN/06/961], GR41350-1. There might have been a problem(s) before we obtained these antibodies.  I would like to mention that this is our first problem with Abcam antibodies, which generally are of very good quality. LOT NUMBER gr36421-2 (We have tried another prior to this with the same result. We bought the third, which we are going to try soon) DESCRIPTION OF THE PROBLEM No band at all SAMPLE •Species: Human •What’s cell line or tissue: human saliva PRIMARY ANTIBODY •Species: mouse •Reacts against: Human, Mouse, Monkey MUC5B •At what dilution(s) have you tested this antibody:1:500 (up to 100) Incubation time: 1h  (ON included)•Incubation temperature: RT (for ON, at 4C) •What washing steps were done: 10mins,3 times DETECTION METHOD: ECl+ POSITIVE AND NEGATIVE CONTROLS USED no but other Polyclonal Ab  (Abnova) works fine (see attached fig). ANTIBODY STORAGE CONDITIONS -20℃ SAMPLE PREPARATION •What lysis buffer was used: commercial •What protease inhibitors were used: Loche mix •What loading buffer was used: 5x loading buffer(include Bromophenol blue +/- DTT) •Phosphatase inhibitors: none •Did you heat the samples: temperature and time: 95℃,5mins AMOUNT OF PROTEIN LOADED 20μg ELECTROPHORESIS/GEL CONDITIONS •Reducing or non reducing gel: reducing gel •Reducing agent: DTT •Gel percentage : 5-20% TRANSFER AND BLOCKING CONDITIONS •Transfer conditions: (Type of membrane, Protein transfer verified):PVDF •Buffer:1x TBST •Blocking agent: milk,  what percentage:5% milk •Incubation time:60mins •Incubation temperature:room temp. SECONDARY ANTIBODY •Species: sheep •Reacts against: human IgG •At what dilution(s) have you tested this antibody: 1:200 •Incubation time: 1h at RT •Wash steps: 10mins,3 times •Fluorochrome or enzyme conjugate:none •Do you know whether the problems you are experiencing come from the secondary? never HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? >6 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes!  

Read More
Answer

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!  

Read More

Question
Answer

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. 

Read More

Answer

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.

Read More

Question

LOT NUMBER gr36421-2 ORDER NUMBER 925356 DESCRIPTION OF THE PROBLEM Non-specific band SAMPLE •Species: mouse •What’s cell line or tissue: saliva •Cell extract or Nuclear extract: none •Purified protein or Recombinant protein: none PRIMARY ANTIBODY •Species:mouse •Reacts against: Human, Mouse, Monkey MUC5B •At what dilution(s) have you tested this antibody:1:500 •What dilution buffer was used: 0.5% blocking buffer •Incubation time: 2 O/N •Incubation temperature: 4℃ •What washing steps were done: 10mins,3 times DETECTION METHOD ECl, ECl+, other detection method: Western Lightning Chemiluminescence Reagent Plus (PerkinElmer Life Sciences, Inc.) POSITIVE AND NEGATIVE CONTROLS USED no ANTIBODY STORAGE CONDITIONS -20℃ SAMPLE PREPARATION •What lysis buffer was used: none •What protease inhibitors were used: none •What loading buffer was used: 4x loading buffer(include Bromophenol blue 、2-β-Mecapitalethanol) •Phosphatase inhibitors: none •Did you heat the samples: temperature and time: 95℃,5mins AMOUNT OF PROTEIN LOADED 10μg ELECTROPHORESIS/GEL CONDITIONS •Reducing or non reducing gel: reducing gel •Reducing agent: 2-β-Mecapitalethanol •Gel percentage : 10% TRANSFER AND BLOCKING CONDITIONS •Transfer conditions: (Type of membrane, Protein transfer verified):PVDF •Buffer:1x TBST •Blocking agent: milk, BSA, serum, what percentage:4% milk •Incubation time:60mins •Incubation temperature:room temp. SECONDARY ANTIBODY •Species:goat •Reacts against: mouse IgG •At what dilution(s) have you tested this antibody: 1:5000 •Incubation time: O/N •Wash steps: 10mins,3 times •Fluorochrome or enzyme conjugate:none •Do you know whether the problems you are experiencing come from the secondary? none HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 2 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? Modify the concentration of 2nd antibody(1:5000 and 1:2000)

Read More
Answer

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.

Read More
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Asthma lung)
Specification
Asthma lung
Fixative
Formaldehyde
Permeabilization
No

Dr. Francois Daubeuf

Verified customer

Submitted Apr 27 2011

1-10 of 11 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com

Sign up