Anti-CXCR4 antibody (ab2074)

Inquiry: I have a couple of questions regarding the CXCR4 antibodies for use on mice tissue. You have 2 products: ab2074 and ab7199. What is the difference between the two and which seems to be more effective (i.e. for immunoblotting and IHC on paraffin embedded tissues)? Also, when I gather tissues from mice, I would like to include an internal positive control. Which tissue would you recommend that can be used as a positive control for CXCR4 immunohistochemistry? Thanks.

ANSWER:

Thank you for your inquiry.

We would guarantee both ab2074 and ab7199 to work in mouse samples in IHC-P and WB. Both are purified and are 1mg/ml. The ab2074 is raised against a human peptide sequence and the ab199 is raised against the full length rat recombinant protein. Please note that ab2074 has a recommended specific protocol:

http://www.abcam.com/assets/popups/popup_datasheet_protocols_printer_friendly.cfm?pid=901&intAbId=2074

Oftentimes customers will choose an antibody that has the most references and Abreviews. Out of the two, ab2074 ismore heavily referenced and we have more IHC-P and WB data.

http://www.abcam.com/CXCR4-antibody-ab2074-references.html

http://www.abcam.com/CXCR4-antibody-ab2074-reviews.html

As for a positive control, we would recommend mouse bone marrow or skeletal muscle. Swiss Prot lists tissue specificity as: Lymphocytes, macrophages, neutrophils, microglial cells and astrocytes. Found in spleen, thymus, bone marrow, lymph nodes and, at lower levels in brain, small intestine, stomach and kidney. CXCR4-A is predominant in all tissues tested. During embryonic development, high levels are detected in the endothelium of developing blood vessels and in many regions of the developing brain including the olfactory epithelium, olfactory bulb, hippocampus, cerebellum and spinal cord.

http://www.uniprot.org/uniprot/P70658

In case you are interested, we sell mouse skeletal muscle slides as ab4609.

http://www.abcam.com/Skeletal-muscle-mouse-normal-tissue-slides-ab4609.html

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

I am writing to ask if it would be possible to obtain a small sample of your Rabbit anti-Cxcr4 antibody (Product code ab2074). We would like to use the antibody to stain brain tissues - fixed, frozen, cyosections using immunofluorescence methods. We'd like to ensure that the product will work for our applications prior to purchasing an entire lot of the antibody.

ANSWER:

Thank you for contacting us.

While we do not provide free samples of any of our products, we will cover them under our Abpromise guarantee. If you are unsatisfied with your results and have been using this antibody on lightly fixed frozen sections from human, mouse, or rat, we will be happy to refund or replace your purchase for you.

If you are interested in using this antibody on perfused-fixed frozen sections (IHC-FoFr), or on an untested species, you would qualify for our 100% testing discount. This program is basically a buy one, get one free. If you purchase the antibody for full price and submit an Abreview with your protocol and results, you will get a discount code for a free primary antibody on a future order.

http://www.abcam.com/collaborationdiscount

I hope this helps, please let me know if you are interested in our testing discount program or if you need any additional information or assistance.

I safely received the ab5506 antibody and tried it out this weekend. I used 5% goat serum as a blocking agent this time. Unfortunately, it still produced no staining in either normal or tumour prostate. I have also tried Lrgi1 and Cxcr4 antibodies (ab2074 & ab36707) without antigen retrieval. To my surprise the staining worked, but it was still nuclear instead of predicted membrane localisation. So no improvement over the protocol with antigen retrieval. I think we'll have to go with the refund option.

ANSWER:

Thank you for you reply.

Your credit note IDs are ***** & *********.

I am sorry that these antibodies did not perform as stated on the datasheet, I have asked our Finance department to issue a credit note for you for all of these antibodies (ab2074, ab5506, ab32392, ab39707, ab51317, ab52971 and ab65006).

The credit notes may be used in one of the following ways:

(1) Redeemed against the original invoice if this hasn't already been paid.
(2) Held on the account for use against a future order.
(3) A full refund can be offered where no other invoices are outstanding.

Please contact your Finance department to confirm how you would like the credit note to be used and ensure it is not redeemed without your knowledge.

To specifically receive a refund please ask your Finance department to contact our Finance department at creditcontrol@abcam.com or by telephone using the information at the “Contact Us” link in the top right corner of our website.

The credit note ID is for your reference only, please refer to the credit note ID in any correspondence with our accounting department. We will send you the completed credit note by email or postal mail with the actual credit note number which will start with the letters CGB.

I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service should you require further expert advice

We've purchased a number of antibodies from AbCam over the course of the past 6 months and we're struggling to get them to work properly. We therefore wonder if you could provide us with some advice to improve their performance. The antibodies in question and their respective dilutions are:
CXCR4 - ab2074 (1:100)
EpCam - ab32392 (1:250)
LRIG1 - ab36707 (1:200)
Integrin beta1 - ab52971 (1:250)
TACD2 - ab65006 (1:200)
c-Kit - ab5506 (1:100)
Sca1 - ab51317 (1:100)
Our samples are murine tumour and normal tissue from Skin, Mammary and Prostate (only Prostate for the last three antibodies). All samples are Formalin fixed and Paraffin embedded. We use a generic immunohistochemistry protocol. Antigen retrieval is performed by heating slides in Dako Citrate buffer at 100C for 20 min. TBS+Tween20 is used as washing solution. The blocking agent (for rabbit antibodies) is 10% normal goat serum. Antibody binding is detected by anti-rabbit HRP-conjugated secondary antibody from Envision kit with subsequent visualisation with DAB reagent. Rabbit anti-rat biotinylated secondary in 10% normal rabbit serum was used to detect Sca1 primary followed by ABC kit (vectastatin) and DAB treatment.

The main problems we've encountered with the named antibodies are the lack of positive staining (for c-Kit and Sca1 antibodies) and non-specific staining (for the rest). CXCR4, LRIG1, EpCam, Integrin beta 1 and TACD2 all displayed nuclear staining of various intensity instead of expected membrane-associated staining pattern.

We would therefore appreciate any suggestions on potential ways to improve the staining. Please, let me know if you would like me to send you detailed protocols and/or images of the stained tissue.

ANSWER:

Thank you for contacting Abcam.

I am sorry about all of the problems you have been having with these antibodies.

It would greatly be appreciated if you could send me some of the images that you are getting using these antibodies and also could you answer the questions below, so that I can understand your protocol a little more:

1 - How long do you block your samples for?

2 - How long do you incubate your primary antibodies for?

3 - Do you use a hydrogen peroxidase step?

4 - Are your samples permeabilised? If so with what percentage of detergent and for how long?

5 - Have you tried testing any of these antibodies without antigen retrieval, as for ab2074 & ab36707 at least, we have some information that suggests antigen retrieval is not necessary.

Once again, I am sorry about all the problems you have been and having and I hope to able to help you resolve this issue.

I look forward to your reply.

Could this antibody work in a neutralising assay?

ANSWER:

DISCOUNT CODE: ***
Expiration date: July 3, 2012

I am very pleased to hear you would like to accept our offer and testab2074 in a neutralising assay. This code will give you 1 freeprimary antibodybefore the expiration date. To redeem this offer, please submit an Abreview for ab2074 in a neutralising assayand include this 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.

The code will be active once the Abreview has 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.