Anti-CXCR4 antibody - Hematopoietic Stem Cell Marker (ab1670)

Would like a control antibody to use with bone marrow samples.

ANSWER:

Thank you for your call today and for your questions about our antibodies to use in IHC.

Theearly hematopoieticsurface marker CD34 is present in bone marrow, as shown on the Human Protein Atlas linkedbelow-

http://www.proteinatlas.org/ENSG00000174059/normal/bone+marrow

We have a CD34 antibody that is known to react with mouse paraffin-embedded or frozen tissue sections, ab8158-

http://www.abcam.com/CD34-antibody-MEC-147-Hematopoietic-Stem-Cell-Marker-ab8158.html

I've also found on the Human Protein Atlas that bone alkaline phosphatase is present in the bone marrow-

http://www.proteinatlas.org/ENSG00000162551/normal/bone+marrow

The bone ALP antibody that we discussed was ab108337-

http://www.abcam.com/Bone-Alkaline-Phosphatase-antibody-EPR4477-ab108337.html

One other option in a neutrophil elastase antibody, such as ab68672, which works with murine paraffin-embedded or frozen tissue-

http://www.abcam.com/Neutrophil-Elastase-antibody-ab68672.html

Neutrophil elastase is also expressed in the bone marrow-

http://www.proteinatlas.org/ENSG00000197561/normal/bone+marrow

I additionally recommend reviewing the literature to confirm that these markersare present for your specific murine time points.

Since ab68672 and ab108337 are rabbit polyclonal primary antibodies, they can be used with our rabbit specific IHC kits such as ab80437, which contains all reagents needed for the IHC staining--

http://www.abcam.com/EXPOSE-Rabbit-specific-HRP-DAB-detection-IHC-kit-ab80437.html

If you prefer to use a fluorescent secondary antibody, we have several pre-adsorbed anti-rabbit secondary antibodies such asab7086-

http://www.abcam.com/Rabbit-IgG-secondary-antibody-H-L-Pre-Adsorbed-ab7086.html

The pre-adsorption treatment will prevent cross-reaction with the mouse tissue that can create background. As ab8158 was raised in a rat, we have several pre-adsorbed anti-rat secondary antibodies such as ab7100-

http://www.abcam.com/Rabbit-polyclonal-Secondary-Antibody-to-Rat-IgG-H-L-FITC-pre-adsorbed-ab7100.html

I hope this information will be useful, but if you have any questions or if there is anything else that you need for your research, please let me know and I'll be happy to help.

I am in need of a CXCR4 antibody that can used for immunohistochemistry in formalin fixed paraffin embedded rat tissue. You have several CXCR4 antibodies (ab13316, ab1670, ab2074, ab1671). Are any of these antibodies compatible with my tissue?

Thanks for your help,

ANSWER:

Thank you for your enquiry. Unfortunately we don't currently have an antibody that has been tested for cross-reactivity with rat and tested for application in IHC-P. Ab2074 and Ab1670 cross-react with mouse and work in IHC-P. You may want to see what the homology is between mouse and rat CXCR4 - one of these antibodies may be suitable for you.

one of our customers uses you CXCR4 antibody #AB1670 for staining of FFPE sections. He used standard antigen demasking using citrate buffer, but observed unspecific background and a weak and diffuse staining. So the question is, what kind of pretreatment do you recommend (a protocol would be best)?

ANSWER:

Here is a suggested protocol from the originator:

Immunohistochemistry on Paraffin sections:

Preparation of slides:

Place slides in 60 C oven overnight after cutting. Then follow the procedure below:

1) xylene 2x10min

2) 100% alcohol 2x10min

3) 95% alcohol 5 min

4) 80% alcohol 5 min

5) rinse in distilled water 2x2 min

6) antigen retrieval if necessary

7) Immunohistochemistry

Antigen Retrieval: Sections were treated with antigen retrieval solution (Dako # S-1700, pH 6.0 or #S-3308, pH 8.0) and steamed for 20 minutes followed by cooling for 20 minutes.

Immunohistochemistry: Block the endogenous peroxide by incubating the sections with 3% H2O2 for 15 minutes at room temperature (RT). Wash the sections with PBS and incubate with 2% normal rabbit serum at room temperature to block the non-specific binding sites. Wash the section with PBS and incubate with primary antibody (anti-chemokine receptor) at recommended dilution for 60 minutes at RT. Wash the slides and incubate at RT with biotinylated rabbit anti-goat IgG H+L (Vector Labs) at 1-200 dilution for 30 minutes. Wash the slides and incubate with HRP-Streptavidin (Vector Labs) 1:400 dilution for 30 minutes. Wash the slides and incubate with DAB (sigma) for 5-7 minutes at RT. Wash slides with PBS and counter stain with hematoxylin (DAKO).

Dear Sir

Regarding #AB1670. Please let me know the molecular weight of band recognized by this antibody. I look forward to receiving your early reply. Best regards

ANSWER:

Thank you for your enquiry and your interest in our products. We would like to confirm that this antibody should recognize a band at approximately 41 kDa.

I have been looking on your website for the following antibodies:

(i) anti-mouse CCR7, application FACS, preferably conjugated to something like FITC or PE (ii) anti-mouse CXCR4, ditto.

However goat or rabbit polyclonals will do. The important thing is recognition of native protein i.e. on the cell surface, which of course sometimes means that abs that are reportedly suitable for Westerns are no good.

Any suggestions? I looked them up on your site but couldn't find any that seemed to be specifically tested for FACS and I wondered if you might be able to nudge me in the direction of some useful ones based on your knowledge of the tests these things go through. Specifically, I thought I'd try ab1657 and ab1670.

ANSWER:

Thank you very much for your enquiry. The only CCR7 that we currently have is ab1657 - Goat polyclonal to CCR7. Of the CXCR4 antibodies that we have, ab1670 - Goat polyclonal to CXCR4 - seems to be the best choice for you as the immunogen is mouse. However, neither of these antibodies have been tested for application yet in FACS so we can't provide any guarantees that they will indeed work. If you decide to go ahead and purchase either of these products, please let us know how you get on and in return we will forward a reward of your choice, typically an Amazon gift voucher. If you have any more questions, please contact us again.