Anti-PMCA1抗体(ab3528)

Thank you for taking time to provide me with that extra information. It has been very helpful.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would like to offer some suggestions to help optimize the results from ab3528.

1. As the target it a membrane bound protein I would suggest using a milder detergent for permiabilisation as Triton X may be too harsh. A detergent such as Tween 20, Saponin or Digitonin may be preferential as these give large enough pores for antibodies to go through without dissolving plasma membrane. It may also be worth trying a different blocking agent, such as 5% milk or BSA.

2. With the Western blot, I would suggest reducing the amount of protein currently loaded to 20 ug. I would also suggest reducing the heating step to 10 minutes at 70°C as this can reduce aggregation, especially when targeting membrane proteins such as PMCA1. I think it would also be worthwhile trying an alternative lysis buffer such as RIPA in order to make sure the protein is well solubilised. As with the ICC experiment, I would also try a different blocking buffer to see if the results are affected, such as 5% milk. This can have a surprisingly stark effect with some antibodies, as illustrated with the anti-GAPDH ab9385:

https://www.abcam.com/ab9385

Should the suggestions not improve the results, please do let me know.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

Thank you for contacting us and for sharing the problems you have been experiencing with the Anti-PMCA1 antibody (ab3528). I am sorry to hear you are experiencing difficulties with this product. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I have a few further questions if you wouldn't mind. This should allow me to more fully understand what may be contributing to these problems.

1. When performing the ICC experiment, was any blocking used, if so what? Also, was any permiabilisation performed?

2. When performing the Western blot with the antibody, could you describe how the samples were prepared? What lysis buffer was used, were any protease inhibitors used? Also, at what dilution was the primary antibody used in?

3. Have the secondary antibodies used been used with any other primary antibodies previously with no problem?

4. Do you have the lot number used (or the order, or the approximate date of the order and the delivery address)?

With this information I hope to be able to understand more fully what may be contributing to these problems.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.