Anti-C3 antibody [11H9] (ab11862)

Product code: 19187
Inquiry: Dear Support-Team, I would need to immunostain mouse/rat C3 on mouse/rat paraffin sections and thus looking for an appropriate antibody in IHC-P. Has ab19187 been tested for IHC-P as well? Given that it was raised by immunizing with purified mouse C3, it should be able to detect mouse C3. Thank you for your information. Kind regards,

ANSWER:

Thank you for your enquiry and your interest in our products.
I have checked this product (ab19187: Anti-C3 antibody) for you and according to the on-line datasheet it has only been tested in Immunoelectrophoresis.
However, we have an alternative antibody in our catalogue ab11862 which recognizes murine C3 and has been characterized for IHC-P (immunohistochemistry on formalin-fixed paraffin-embedded tissue sections).
If you need any further assistance, please do not hesitate to contact me.

Dear technical team,

Our customer is looking for C3 antibody which can detect onlybeta chain.
I'm not sure about alpha or beta subunit so, please recommend antibody for our customer.
His sample is human and mouse and is going to test WB with C3 antibody.
Please infrom me.

Best regard,

ANSWER:

Thank you for your enquiry.

I am sorry to confirm that the C3 antibodies that detect the C3beta subunit have not been tested in WB, human and mouse.

We do have ab11862 Anti-C3 antibody [11H9] which has been tested in WB and human and detects C3, C3b, iC3b and C3b. It should not detect C3 alpha subunit on its own.

I have checked the alignment of the immunogen with the mouse protein, and this is 77%. We recommend that alignment should be over 85% to predict that an antibody will detect in a different species. 77% is fairly high and I would suggest there is still a possibility of some crossreactivity with mouse. However, I am sorry we are not able to guarantee this without testing.

If the customer would like to try the antibody in mouse as part of our testing discount program, please do not hesitate to let me know and I will be pleased to arrange a discount code for them.

I hope this will be helpful. If you have any further questions, please do not hesitate to contact me.

I would like to test these antibodies in rabbit in IHC-P, which has not yet been tested.

ANSWER:

I am very pleased to hear you would like to accept our offer and test ab6678 in rabbit, ab8878 in IHC-P, and ab11862 in rabbit. These codes will give you 1 free primary antibody per code before the expiration date. To redeem this offer, please submit an Abreview for each antibody and include the code in the “Additional Comments” section so we know the Abreview is for this promotion. For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews.

Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The codes will be active once the Abreviews have been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code.

Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

What are the molecular weights of the proteins this antibody cross-reacts with?

ANSWER:

Thank you for your enquiry.

The mature protein C3 has a molecular weight of approximately 190 kDa. The complement factor C3 consists of an alpha- and a beta-chain, linked by disulfide bond. C3 convertase activates C3 by cleaving the alpha chain, releasing C3a anaphylotoxin and generating C3b (alpha chain and beta chain). C3b has a molecular weight of approximately 185 kDa. C3b is rapidly split in two positions by factor I and a cofactor to form iC3b (inactivated C3b) and C3f which is released. iC3b has a molecular weight of approximately 182 kDa.

We hope this information will be helpful to you. If you have further questions please do not hesitate to contact us.

This antibody has not worked in a western blot. The blot was tested as good using a polclonal antibody against the same protein. The serum was prepared as follows: blood was drawn by cardiac puncture (mice under anesthesia) and then placed in the"capillary serum separator" (monoject cat # 008018), let stand at RT for 45 min and then spin at 1200 g. the sup was aliquoted and stored at -80. For the western I used serum and serum that was activated; the activated serum contains cleaved C3 and a band at 46 and 35 is expected if you have a polyclonal antibody (which I saw with the polyclonal).

The lot number from your ab is 152789. Have you had other complaints about this lot number/ antibody in mouse serum? Your website states that it reacts with mouse protein, what is the positive control sample that was used to test this, serum prepared as above? Please provide any additional information about how this antibody was tested.

ANSWER:

Thank you for your patience and your e-mail regarding ab11862 monoclonal antibody to mouse C3.

As a positive control affinity-purified mouse C3 was used. For the purification of mouse C3 purified antibodies were coupled to CNBr-activated Sepharose 4B. EDTA-plasma from heparinized C57BL/6 mice was applied to the column in the presence of protease inhibitor(PMSF, 10 uM). Bound complement components were eluted with 50 mM diethylamine (pH 11.5) and immediately neutralized with 1/10 volume of 500 mM NaH2PO4. Contamination by rat mAbs were removed from the eluates by protein G sepharose 4 Fast Flow in a batch procedure.

For more information please see: 1) Kremmer, E et al; Monoclonal antibodies to complement complement component without the need of their prior purification. II Antibodies to mouse C3 and C4. Hybridoma 1990, 9: 309. Please note we did not receive any other complaints regarding this lot, however we take this complaint very seriously.

Please contact us again if you require further assistance and do not have success with the antibody.