Anti-CD51 + CD61 antibody [BV3] (ab7166)

Thank you for your reply, that is very interesting. We had selected to use CD51/61 as a marker for angiogenesis, we are looking into the pathological outcome of possible angiogenesis in brain tissue. We had selected the glioblastoma case as a positive control to test the antibody before running it through our cohort. Do you have any recommendations of how we can increase specific staining of the antibody? Do you believe then that staining of blood is just monocytes, we had suspected that it maybe red blood cells.
I look forward to hearing your response
Kind regards

ANSWER:

Thank you for your email.

The sensitivity of staining could be increased by;

- using enhancers; we have DAB enhancer in catalogue, please check the datasheet http://www.abcam.com/DAB-Enhancer-ab675.html
- Using lower dilution of primary or secondary abs without compromising the background
- Incubating primary overnight at 4C and secondary for 2 hours RT
- Ab Optimization; In IHC-P optimization of antibodies is always necessary for finding the right dilution and best antigen retrieval procedure e.g. sometime 5 min microwave give clean staining than 10 min microwave. In my own experience microwave 10 minutes gave cleaner staining than 5 minutes so it is good to check which experimental condition is best suitable in relation to tissue section and antibodies.

RBCs do not express CD51 and CD61 so the antibody should not bind to these cells. To confirm this you may consider dipping the slides in hematoxylin solution, RBCs do not have nucleus so they will not be stained.

The following website is a also a good resource for searching the endothelial targets; http://www.antibodybeyond.com/reviews/tumor-markers/angiogenesis-marker.htm

Let me know what you think.

I hope this information will be helpful. Should you have any question please do not hesitate to contact me.

I have converted images to JPEG hopefully that will help. Please find attached images of glioblastoma case 2 images of microwave retrieval and 2 with pressure cooker, both using EDTA. Tried citrate buffer before and found similar problems. I have checked with the technicians and the case has low post mortem delay and fixation time

Kind regards

ANSWER:

Thank you for your email.

The CD51 and CD61 target is expressed by endothelial cells, megakaryocytes, monocytes, macrophages and Neutophils which is why the staining is more in the blood vessels.

It seems the antibody is binding to the cells where it suppose to bind however the staining is quite faint.

I suppose you are studying Glioblastoma in human brain which involves glial cells. Could I ask the reason of choosing CD51 and CD61 targets.

I will look forward to hearing from you soon.

Antibody giving high background
- Looks like staining not specific to endothelial cells
- Interested in observing Gleoblastoma
- Human brain tissue sections (post mortem)
- Citrate buffer, EDTA buffer usedwith 10 minutes heating in microwave and pressure cooker.
- Ab dilution used 1/100, 1/500 and 1/1000 no improvement

ANSWER:

Thank you for contacting us.

As discussed on phone please attach images and send it to us.

We will look forward to hearing from you soon.

The customer has submittedthe Abreview, has it been accepted?

The email approval received by the customer mentions abreview points and not FOC antibody, I assume it's only the automatic reply. Could you please check if the customer is entitled for a FOC antibody.

Thank you for your kind help.

Regards,

ANSWER:

Thank you for your message.

Yes, I can confirm that this Abreview has been published. Please thank the customer for their valuable feedback.

You can now use the discount code when the customer places an order for their next primary antibody. The usual procedure is to include the code when you place the order for thefree of charge antibody the customer would like to receive.

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

I have used both of these antibodies for flow cytometry and immunohistochemistry. I saw that both can be used for immunoprecipitation. I would like to know if you all have an idea whether these antibodies may also work for western blots or you've tried them for WBs and they didn't work?

ANSWER:

Thank you for contacting Abcam. I have checked our notes and it appears that ab7166 is unsuitable for western blot, while ab24694 will most probably work for western blotting, its just that we have not tested it in that application. Please let me know if there is anything else I can help you with.