Recombinant Anti-LEF1 antibody [EP2030Y] (ab53293)

Thank you for your inquiry.
The exact immunogen sequence is proprietary information and I unfortunately do not have access to it.
But the lab confirmed that all isoforms should be recognized by ab53293 since all isoforms contain the immunogen sequence with a 100% homology.
This has not been tested experimentally, but a cross-reaction is very likely due to the perfect homology.
I hope this information is helpful and wish you good luck with your research.

Thank you for taking the time to contact us. I am sorry to hearthe customer has some concerns regarding the results from this antibody.

I would like to reassure you that this antibody is tested and covered by our 6 month guarantee for WB and human samples. In the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund.

With regards to the band size, according to the SwissProt/Uniprot protein database site, there are several smaller isoforms of LEF1, but no larger ones at 70 kDa. Could you confirm the customer is reducing and denaturing the samples and running them on a reducing gel? This will ensure the protein is in the correct conformation to run at the correct molecular weight.

I would like to investigate this particular case further for you, and also obtain more information for our quality monitoring records. In order to proceed with this, I have enclosed a technical questionnaire below. I would appreciate if you could complete this. It will help you put the information we require together very easily.

I would appreciate if you could also provide an image (including molecular weight markers)which would help us to assess the results.

Thank you for your time and cooperation. We look forward to receiving the completed questionnaire and image.

Order Details
Antibody code:

Problem
Choose: Non-specific band Multiple bands No signal or weak signal High background

Lot number

Purchase order number
or preferably Abcam order number:



General Information
Antibody storage conditions (temperature/reconstitution etc)


Description of the problem (high background, wrong band size, more bands, no band etc.)


Sample (Species/Cell extract/Nuclear extract/Purified protein/Recombinant protein etc.)


Sample preparation (Buffer/Protease inhibitors/Heating sample etc.)


Amount of protein loaded


Electrophoresis/Gel conditions (Reducing or Non-reducing gel, % of the gel etc.)


Transfer and blocking conditions (Buffer/time period, Blocking agent etc.)


Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)


Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)


Detection method (ECL, ECLPlus etc.)


Positive and negative controls used (please specify)



Optimization attempts (problem solving)
How many times have you tried the Western?



Have you run a "No Primary" control?
Yes No

Do you obtain the same results every time?
Yes No
e.g. are the background bands always in the same place?


What steps have you altered?


Additional Notes:


Image:
We would appreciate if you are able to provide an image (including molecular weight markers) which would help us to asess the results.

Thank you for contacting Abcam.

I am sorry that you are having problems with ab53293 in western blot. If you are looking for a positive control then I would recommend ab7899:

https://www.abcam.com/Jurkat-Human-T-cell-lymphoblast-like-cell-line-Whole-Cell-Lysate-ab7899.html



Also I would be very happy to try and help you resolve your western blot issues, if you could answer the questions below, I will see if thereare any protocol changes that I can suggest:

1 - What sample type are you using (including species)?

2 - Are you seeing any bands, no bands etc?

3 - How much sample are you loading?

4 - What type of blocking agent are you using (eg milk, BSA etc)?

5 - What primary antibody concentrations have you tried and how long do you incubate for?

I look forward to your reply and helping you resolve this issue.