Anti-Aggrecan ARGxx antibody [BC-3] (ab3773)

TSC - was cultured in serum?
is the Ab available in purified form?

ANSWER:

TSC - was cultured in serum?
A: Serum was used during culture.

is the Ab available in purified form?
A: We do not routinely purify the antibody and so cannot give an answer to the second question. However, it could be purified.

I hope this is helpful. Please contact us again if you have any further questions.

Dear technical team, Our customer would like to purchase 5 antibodies from abcam and wants to check control tissue. Pleaselet me knowabcam's product information. ab66025 ab102711 ab3773 ab2426 ab58632 Best regards,

ANSWER:

Thank you for your inquiry. Please find a list of possible positive controls below: ab2426: skin, lymph nodes Click here (or use the following: http://www.abcam.com/index.html?datasheet=2426). ab3773: cartilage samples Click here (or use the following: http://www.abcam.com/index.html?datasheet=3773). ab58632: Type X collagen is a product of hyperthrophic chondrotocytes and has been localized to presumptive mineralization zones of hyaline cartilage. Click here (or use the following: http://www.abcam.com/index.html?datasheet=58632). ab66025: The recommended positive control tissues are rat head tissue, rat embryo tissue, human osteosarcoma tissue, human glioma tissue, human intestinal cancer tissue. Click here (or use the following: http://www.abcam.com/index.html?datasheet=66025). ab102711: Specifically expressed in osteoblasts Click here (or use the following: http://www.abcam.com/index.html?datasheet=102711). I hope this information is helpful for your customer. I wish them good luck with their experiments.

Which antibody is most suitable for detecting full length aggregan protein in goat tissue?

ANSWER:

I have been informed of the following information which I hope will help you:

Ab3773: this should be reactive with goat since this antibody recognises a sequence ARG... that is conserved in all species Guinea Pig/rat to human but not mouse for some reason where the sequence is ALG.... This antibody will not recognise the sequence ...ARG.. if it is in the non-cleaved intact aggrecan protein core; i.e. it will only recognise the aggrecanase generated neoepitope ARG...

Ab3778: this antibody recognises a linear sequence ..EPEEP... that is present in the IGD protein core of human and bovine aggrecan but not pig and several other animals; it seems it is not present in goat protein after BLASTing the sequence in goat. This antibody recognises the linear epitope ...EPEEP... in both intact and matrix protease degraded aggrecan from human and bovine cartilage.

Please do not hesitate to contact me if I can be of further help,

I want to buy a mouse monoclonal antibody to aggrecan ARGxxx Cat no. ab3773. I wang to use it for immunohistochemistry for aggrecan of rattus chondrocyte. I want to konw whether I must treat the chondrocytes with digestion mixture ( Chondroitnase ABC Sigma, Keratanase Seikagaku and keratanase II Seikagaku ) as recommended before immunohistochemistry or western-blot.

ANSWER:

Thank you for your enquiry.

The following reference gives details of the protocol used with this antibody in immunohistochemistry: Roberts S et al. Matrix metalloproteinases and aggrecanase: their role in disorders of the human intervertebral disc. Spine 25:3005-13 (2000). PubMed: 11145811

I have added the relevant protocol to our online datasheet for your convenience and include below the following publications which have all used this product.

This product has been used in:

Karsdal MA et al. Induction of increased cAMP levels in articular chondrocytes blocks matrix metalloproteinase-mediated cartilage degradation, but not aggrecanase-mediated cartilage degradation. Arthritis Rheum 56:1549-58 (2007). PubMed: 17469134

Little CB et al. Matrix metalloproteinases are involved in C-terminal and interglobular domain processing of cartilage aggrecan in late stage cartilage degradation. Matrix Biol 21:271-88 (2002). PubMed: 12009333

Flannery CR et al. Autocatalytic cleavage of ADAMTS-4 (Aggrecanase-1) reveals multiple glycosaminoglycan-binding sites. J Biol Chem 277:42775-80 (2002). PubMed: 12202483

Malfait AM et al. Inhibition of ADAM-TS4 and ADAM-TS5 prevents aggrecan degradation in osteoarthritic cartilage. J Biol Chem 277:22201-8 (2002). PubMed: 11956193

Roberts S et al. Matrix metalloproteinases and aggrecanase: their role in disorders of the human intervertebral disc. Spine 25:3005-13 (2000). PubMed: 11145811

Caterson B et al. Mechanisms involved in cartilage proteoglycan catabolism. Matrix Biol 19:333-44 (2000). PubMed: 10963994

Billington CJ et al. An aggrecan-degrading activity associated with chondrocyte membranes. Biochem J 336 ( Pt 1):207-12 (1998). PubMed: 9806902

Buttle DJ et al. "Aggrecanase" activity is implicated in tumour necrosis factor alpha mediated cartilage aggrecan breakdown but is not detected by an in vitro assay. Mol Pathol 50:153-9 (1997). PubMed: 9292151

Hughes CE et al. Monoclonal antibodies that specifically recognize neoepitope sequences generated by 'aggrecanase' and matrix metalloproteinase cleavage of aggrecan: application to catabolism in situ and in vitro. Biochem J 305 ( Pt 3):799-804 (1995). PubMed: 7531436

This clone has been used in:

Molecular Approaches to Osteoarthritis Research. Abstracts of the Kennedy Institute of Rheumatology Symposium, London, United Kingdom, April 18-20, 2004. Scand J Rheumatol Suppl :1-41 (2005). PubMed: 15864865

Roberts S et al. Matrix turnover in human cartilage repair tissue in autologous chondrocyte implantation. Arthritis Rheum 44:2586-98 (2001). PubMed: 11710715

Aplin JD et al. Human endometrial MUC1 carries keratan sulfate: characteristic glycoforms in the luminal epithelium at receptivity. Glycobiology 8:269-76 (1998). PubMed: 9451036

Can you tell me the molecular weigth for the full-length protein and also the protein after it has been degraded? I am using rabbit and guinea pig tissue.

ANSWER:

Thank you for your patience. I can see that my colleague has also written to you and referred you to the paper in which the antibody was tested, i.e. Little CB et al., Matrix Biol 21:271-88 (2002). In summary the antibody detects a variety of epitopes between 50 and 250 kDa.

I also contacted the originator of the antibody. Using ab3773 in Western blots can provide a wide range (of MW) of bands that are immuno-positive for the BC-3 neoepitope (i.e. ARG... as a result of IGD aggrecanase cleavage). Thus, if you are looking at media from cartilage explant cultures exposed to cytokines such as IL-1 you will find a wide range of MW product usually from 50kD - 250kD. However, if you use cartilage from young animals this can give mainly large MW bands and if the aggrecanase activity has been low you may see only bands at 40/50 - 90kD.

I hope this additional information helps you.