Anti-EGFR antibody (ab2430)

Dear technical team,

Our customer would like toreceive CoC of ab2430 with lot#GR69660-1.
Please inform me for our customer.

Best regard,

ANSWER:

Thank you for contacting us and sorry for the delay in getting back to you.

Please find the Certificate of compliance attached as requested.

If you have any further questions please do not hesitate to contact us again.

Dear technical team,

Our customer would like to receive CoC of ab2430 (lot#GR69660-1).

Please send me the file.

ANSWER:

Thank you for contacting us.

I have attached the CoC for AB2430 as you have requested.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Thank you for your below reply and for your support.



Kindly find below in blue the customer’s reply to your inquiries concerning this antibody:



1. What is the lysis buffer that you have used? The lysis buffer used is laemmli buffer.

2. Have you reduced and denatured your samples? Which method/buffers did you use? The samples were denatured and reduced by heating at 95 degree Celsius for 10 minutes and then adding Beta-mercaptoethanol (10% of total volume).

3. How much total protein did you use per well? 25 microgrof each sample is added per well.

4. What membrane type did you transfer to? Nitrocellulose membrane is used for transfer (Amercham).

5. What blocking steps did you use? 5% milk (zero fat) is used for blocking (2.5 grams of this powdered milk is dissolved in 50 ml wash buffer: TBS tween 20).

6. What dilutions of this product did you use? How long and at what temperature was this antibody incubated? The primary antibody was used at 1/ 1000 dilution (for EGFR antibody ) shaking at room temperature for 2 hours.

7. What was the secondary antibody that you have used? How long and at what temperature was this antibody incubated? Anti-rabbit was used as secondary antibody. Dilution: 1/ 2000 at room temperature shakingfor 2 hours.

8. Have you run a no primary antibody control blot? What were the results? yes, theno primary antibody control shows no band.

9. What is the wash buffer that you use? TBS tween 20 is the wash buffer

ANSWER:

Thank you for providing that protocol information.

Having looked through the Abreviews that are on the website for ab2430, there are 2 protocol variations that I can suggest:

1 - Instead of using a 5% milk block, try using a 5%BSA block.

2 - Incubate the primary antibody in BSA, overnight at 4C, instead of at room temperature.

If these variations do prove to be successful, or if you feel that they will not work, then please let me know and I will be happy to either replace or refund the cost of the antibody. If you do wish to receive a replacement or a refund, then please provide me with the original Abcam order information and also the catalogue number that you would like to receive as a replacement.

Please let me know if there is anything else I can help you with.

We have received the following complaint from a customer who have previously ordered the “ab2430 Anti EGFR antibody”. In the datasheet of this antibody and in all the publications it is mentioned that this protein should appear at 170KDa, but in all his results obtained using different samples the protein appears at 70kDa and not 170kDa.   Kindly find attached to this message the data resulting from using this antibody sent from the customer, kindly advise?

ANSWER:

Thank you for bringing this to our attention. While EGFR has a number of isotypes that appear at 70kDA, this particular product should only recognize the 170kDA isoform. I would appreciate if the customer could provideparticulars of their protocol. This information is important not only in helping us understand the problems but in our internal testing as well.

As always, this product is covered by our Abpromise guarantee.

If we cannot remedy this issue and this is a product that you have purchased within the last six months, we will replace or refund it under our Abpromise guarantee, as you are using it according to specifications listed on our datasheet.

Below is a guide for the type of information which would be most valuable to me in understanding the procedure used:

Could you tell me the lysis buffer that you have used?

Have you reduced and denatured your samples? Which method/buffers did you use?

How much total protein did you use per well?

What membrane type did you transfer to?

What blocking steps did you use?

What dilutions of this product did you use? How long and at what temperature was this antibody incubated?

What was the secondary antibody that you have used? How long and at what temperature was this antibody incubated?

Have you run a no primary antibody control blot? What were the results?

What is the wash buffer that you use?

Could you provide the Abcam order confirmation number or PO number used to purchase this?

Thank you very much for taking the time to answer these questions. Please contact me if you have any questions or concerns.

the order number was: xx
Thanks,

ANSWER:

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with a vial of a new batch with the order number xx.

To check the status of the order please contact our Customer Service team and reference this number.

Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

I wish you the best of luck with your research.