Anti-FOXP3 antibody (ab10563)

Hell


As you know we have ordered a number of anti-human ab from Abcam. The list
is attached...
I would like to request fro your assistance in the following...
We would like to get the information about the recommended diluent for the
ab to be used for IHC with paraffin embedded tissue sections since its known
to affect results at times.
Other information regarding antigen retrieval conditions etc will be
available on the net, but could be refurbished if they are updated.
Equally important is the source of positive and negative controls.
I would especially like to request for slides of both the types of controls
as they are vital to decide/judge the variations in the protocols. I hope
this is done as it will serve us for a long time in future.

ANSWER:

Thank you for contacting us.

I have attached the filled out Exel spreadsheet with all the requested information. To be fair, all the information asked could be answered with a look on the datasheet...

In addition, I had a look through our tissue slices for her IHC-P experiments. Instead of the Tonsils she can use Lymphknot (as we do not have Tonsil slides).

http://www.abcam.com/index.html?datasheet=4350 (or use the following: http://www.abcam.com/index.html?datasheet=4350). PC for ab25101and ab10563

http://www.abcam.com/index.html?datasheet=4348 (or use the following: http://www.abcam.com/index.html?datasheet=4348). PC for ab34843

http://www.abcam.com/index.html?datasheet=4372 (or use the following: http://www.abcam.com/index.html?datasheet=4372). PC for ab37158

Thanks for your kindly reply; this customer replied your question as follow:



1. First antibody: human FOXP3 ab10563
Secondary antibody: anti-rabbit Poly-HRP-IgG
2. The antigen retrieval method and the buffer: Leica Bondmax, EDTA pH=9
3. Was the same procedure used with the two lots? Yes
4. Any troubleshooting tried with protocol? The last primary antibody dilution is 500x. Using
new lot, dilution 200x and 500x, the result is fail. The staining protocols
are the same.



Could you please help this customer to solve the problem?

Thanks for your kindly help

Best regards

ANSWER:

Thank you for contacting us.

Your credit note ID is XXXXX.

I am sorry that this antibody did not perform as stated on the datasheet, I have asked our Finance department to issue a credit note for you. The credit note may be used in one of the following ways:

(1) Redeemed against the original invoice if this hasn't already been paid.
(2) Held on the account for use against a future order.
(3) A full refund can be offered where no other invoices are outstanding.

Please contact your Finance department to confirm how you would like the credit note to be used and ensure it is not redeemed without your knowledge.

To specifically receive a refund please ask your Finance department to contact our Finance department at creditcontrol@abcam.com or by telephone using the information at the “Contact Us” link in the top right corner of our website.

The credit note ID is for your reference only, please refer to the credit note ID in any correspondence with our accounting department. We will send you the completed credit note by email or postal mail with the actual credit note number which will start with the letters CGB.

I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service should you require further expert advice

ich bin an einer immunohistochemischen Färbung von humanem CD8 und humanem FoxP3 (in Mausgewebe) interessiert.



- Idealerweise mit einen rabbit anti CD-8 Primärantikörper

- Und FoxP3 mit Alkalischer Phosphatase (o.a.) gelabeltem Sekundärantikörper, um ggf. später ein doublestaining zu machen



Dazu hätte ich zwei Fragen: Welcher Unterschied besteht zwischen Antikörpern aus Überstanden bzw. aufgereinigten Antikörpern? Gibt es spezielle Vor-oder Nachteile bei den jeweiligen Methoden?



Würden Sie mir bitte ein Angebot mit jeweils passenden Sekundärantikörper machen?

ANSWER:

Vielen Dank für Ihre Anfrage.

Ich habe Ihnen hier vier Antikörper herausgesucht, die wenig bzw. gar keine Kreuzreaktion mit dem Maus-Protein aufweisen, so dass Sie nur das humane Protein im Mausgewebe detektieren sollten.

Drei der vier Antikörper stammen aus dem Kaninchen, falls Sie also eine Doppelfärbung durchführen möchten, wurde ich Vorschlagen, einen der Rabbit Antikörper mit einem ungelabelten Sekundär- Antikörper zu blockieren, und dann den sekundären Antikörper mit einem tertiären Antikörper nachzuweisen: zb. im Falle der Kombination aus ab93278 und ab10563.

Dazu passen wuerden dann die folgenden Antikörper: ab6831 Chicken polyclonal Secondary Antibody to Rabbit IgG - H&L gefolgt von ab97142 Goat polyclonal Secondary Antibody to Chicken IgY - H&L (Alkaline Phosphatase), pre-adsorbed ; und ab7084 Donkey polyclonal Secondary Antibody to Rabbit IgG - H&L (AP), pre-adsorbed.

Oder aber Sie benutzen zwei primäre Antikörper aus zwei verschiedenen Species (ab2481 und ab4055).

Als Sekundär- Antikoerper kommen dann ab7084 Donkey polyclonal Secondary Antibody to Rabbit IgG - H&L (AP), pre-adsorbed; und ab6886 Donkey anti-Goat IgG H&L (AP) secondary antibody.

Bezüglich Ihrer Frage nach dem Aufreinigungsgrad: Der einzige Vorteil eines aufgereinigten Antikörpers ist es, dass Sie eine Isotopenkontrolle berechnen können, da Sie genau wissen, wie viel Antikörper Sie einsetzen. Dies ist aber eigentlich nur in der Flow Zytometrie von Interesse.


Ich hoffe, dies hilft Ihnen weiter. Bitte zögern Sie nicht, sich wieder bei uns zu melden, falls Sie weitere Fragen haben.

Benutzen Sie unsere Produkte? Schicken Sie uns einen Abreview. Verdienen Sie sich eine Belohnung!
http://www.abcam.com/abreviews

Dear abcam technical support team:
This customer already bought ab10563 (Anti-FOXP3 antibody) twice, and she got two different lot numbers as follow:
First time (2010/3/8): Lot: 797994 po: 648851<< Got a good result
This time (2012/3/19): Lot: gr74470-1, po: 1054295 << no signal
This customer thinks this antibody which with lot number gr74470-1 was lost its function already. Therefore, she would like to ask for your help to create a credit note to her, I also attached her experiment step and data in this letter please check this and could you please offer any help to this customer, thanks for your assistance.
Best regards

ANSWER:

Thank you for contacting us.

In order to understand the problem I would like toask following queations;

- In the attached details it is mentioned that Rabbit secondary was used. Could you please double check because rabbit secondary can't used with rabbit primary.

- What was the antigen retrieval method and retrieval buffer used?

- Was the same procedure used with the two lots?

- any troubleshooting tried with protocol?


I will look forward to hearing from you soon.

Please could you send us a replacement vial of ab10563?
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 the order number 1070763.
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.