Anti-Smoothelin antibody [R4A] (ab8969)

No stainingusing a vial of ab8969 on human heart and dog tissues.

ANSWER:

Thank you for your call today and for letting us know about the trouble with ab8969.

As we discussed, I'm sending a free of charge vial from a new lot on the order ***. This should arrive tomorrow.

Please keep me updated about the results with this new vial, and let me know if you have any questions or if there is anything else that we can do for you.

Thank you for your email. We appreciate you bringing this issue to our attention, and yes we would like to be compensated for this product as we are particularly interested in smooth muscle myosin heavy chain 2. We purchased this product in xxx and so almost a year has passed in using this antibody. Several experiments have been carried out by myself and other members of our lab using this antibody and other reagents necessary to the experiments. As you will accept these experiments represent considerable costs to our laboratory, and we would query as to any additional compensation you are willing to provide in this matter. Ideally as well as the refund offered, we would like to have this antibody replaced with the appropriate smooth muscle myosin heavy chain 2 antibody, at no charge to our laboratory.

Additionally, on this same order, we purchased an antibody against smoothelin (ab8969). We have now found this antigen to be expressed in human umbilical vein endothelial cells and in an endothelial cell line. This was unexpected and has made us doubt the specificity of the antibody. We would like to query this matter.

We have purchased several antibodies form Abcam over the years and retain confidence in Abcam products.

ANSWER:

Thank you for your reply.


For ab683, I'd be happy to offer you the following:

Either a refund or credit for ab683 plus another antibody free of charge, or alternatively 2 antibodies of your choice free of charge. Please let me know which option you would prefer.

Currently, we only carry antibodies to smooth muscle myosin heavy chain 1 and 2. We have 7 of those: ab77967, ab82255, ab97539, ab124679, ab81031, ab106919, and ab17811. Please let me know as well if any of these antibodies would be of interest to you.

As for ab8969, please send me an email with more experimental details (such as antibody dilutions, results obtained, detection system used, sample type used (cultured cell or tissue sections), etc.), and I'd be happy to look into this.

I look forward to hear back from you and assist you further.

We are not getting any signal in our Western blots, but had good results with the previous lot using the same protocol and samples.

ANSWER:

Thank you for contacting us.

I am sorry that the product you received did not perform as stated on the datasheet. I have issued a free of charge replacement order with the order number of 199199. You will receive an email confirmation with shipping details shortly.

I apologize for the inconvenience. Please do not hesitate to contact us if you need anything further.

Hello ThereI have a query regarding an antibody that I received from you. It is Ab8969/ Smoothelin antibody, directed against contractile SMC's.I have been trying to optimize this antibody for sometime now - and the pattern of staining that I am getting is not consistant with smooth muscle cells, BUT rather of Trophoblasts (found in Epithelial tissue). The control tissue that I have been using is placenta & tonsil, both containing several vessels and arteries which should contain smoothelin positive SMC's. However in both controls, only trophoblasts stain up, AND extremely specifically. I can send you an image if that would help you.The pathologists suggest that the antibody in the vial is incorrect, and in fact not anti- Smoothelin at all, since there is absolutely no trace of staining in a single artery or contractile vessel. Is this possible???? This is a really important study we are doing in cardiac research, and we would like to demonstrate smoothelin.I would appreciate your assistance in this rather mystifying situation.Many Thanks

ANSWER:

Thank you for your enquiry and your interest in our products.

We would like to ask you please to provide some more information about your IHC staining and fill in this specific IHC Questionnaire. Your answer will help us identify the source of the problem and give you sufficient technical support.

1. Order details: • Batch number: • Abcam order or Purchase order number: • Antibody storage conditions (temperature/reconstitution etc)

2. Please describe the problem (high background, no staining etc).

3. On what material are you testing the antibody in IHC • Species: • Cell Iine or tissue:

4. How did you fix the samples? • Ethanol, methanol: (was it ice cold) • Acetone: (was it ice cold) • Paraformaldehyde (percentage, perfusion fixed on not): • Fixation time: • Fixation temperature:

5. For formalin-fixed paraffin embedded tissue: did you apply antigen retrieval step? • Enzymatic method: • Heat mediated technique: • Other :

6. Blocking steps: • For HRP detection method: did you block endogenous peroxidases: • How did you block the unspecific binding sites: serum, BSA, Milk, other • For how long:

7. Primary antibody • Specification (in which species was it raised against): • At what dilution(s) have you tested this antibody: • What dilution buffer was used: • Incubation time: • Incubation temperature: • What washing steps were done:

8. Secondary antibody • Specification (in which species was it raised against): • What dilution buffer was used: • Incubation time: • Incubation temperature: • What washing steps were done: • Do you know whether the problems you are experiencing come from the secondary?

9. What detection method are you using?

10. Background staining • Please provide an image of your staining

11. Which detection system did you use?

12. Did you apply positive and negative controls along with the samples? Please specify.

13. Optimization attempts • How many times have you tried the IHC? • Do you obtain the same results every time? • What steps have you altered?

We look forward to hearing from you soon.

Can you tell me the concentration of this ab or reccommendations for dilutions to use for immunohistochemistry?

ANSWER:

Ab8969 is in an unpurified form (tissue culture supernatant) and we cannot guarantee the concentrations for unpurified material. When we know what the recommended dilutions are for an antibody we will state them on the datasheet, otherwise the dilutions are assay dependent.