Recombinant full length protein (S. cerevisiae).
Our Abpromise guarantee covers the use of ab4753 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/4000 - 1/8000. Detects a band of approximately 14 kDa. This antibody using the specified conditions may recognize other prominent intrinsic bands (UBLs or their conjugates). Other intrinsic bands are readily detectable in yeast lysates at lower antibody dilutions.|
|IHC-P||Use at an assay dependent concentration. PubMed: 20174468|
|ELISA||1/20000 - 1/100000.|
|Electron Microscopy||Use at an assay dependent concentration. PubMed: 25195618|
Ab4753, generated by immunization with recombinant yeast ATG8 (or Apg 8), was tested by immunoblot with other anti-UBL (Ubiquitin-like modifier) antibodies against E.coli lysates expressing the ATG8-GFP fusion protein (Apg 8-GFP in both panels)). All UBLs possess limited homology to Ubiquitin and to each other, therefore it is important to know the degree of reactivity of each antibody against each UBL.
Panel A shows total protein staining using ponceau.
Panel B shows specific reaction with ATG8 (Apg 8) using a 1:4,000 and 1:8,000 dilution of ab4753 followed by reaction with a 1:15,000 dilution of HRP Goat-a-Rabbit IgG (ab7090). All primary antibodies were diluted in TTBS buffer supplemented with 5% non-fat milk and incubated with the membranes overnight at 4° C. E.coli lysate proteins were separated by SDS-PAGE using a 15% gel.
Similar experiments (data not shown), where other UBL fusion proteins were separated and probed with this antibody showed no reactivity of anti-APG8 (anti-Apg 8) with other UBLs. This data indicates that anti-ATG8 (anti-Apg8)is highly specific and does not cross react with other UBLs. A chemiluminescence system was used for signal detection (Roche). Other detection systems will yield similar results.
Data contributed by M. Malakhov, www.lifesensors.com, personal communication.
This image is courtesy of an anonymous abreview.
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