アブカムでは最適な動作のために Google Chrome など最新ブラウザでの閲覧を推奨します。
We ordered the antibodies anti-GABA transporter 3 (ab431) and the anti-GABA transporter 2 (ab2896). I would like to ask you, which specific size are both detectable in the blott? For example for GAT- 2 in the datasheet of product information there is nothing info about the size, and for GAT-3 in the datasheet says that this antibody reacts with a 74kDa protein from CNS samples (we are doing in rat brain samples), however after perform the western blott, there is no band at this size, and we have a band at 100 kDa and other unspecific bands between 20 and 50.
Asked on Apr 06 2012
Thank you for contacting us.
The GAT3 antibody ab431 detected a band at 74 kDa in CNS samples, as noted on the datasheet. The precursor of the protein, before any post-translation modifications has a predicted molecular weight of about 70 kDa. Glycosylation of the GAT3 is presumed to cause the protein to run in the gel to 74 kDa.
Your result, detection at 100 kDa and 20 and 50, suggests the antibody is detecting something other than GAT3. Degradation of the sample might explain the bands at 20 and 50 kDa, if you are not using protesase inhibitors in your sample lysis buffer, but I have no explanation for the 100 kDa band.
In your reply, could you please include an image of the blot, and a few details of your protocol, such as how much sample you loaded per lane of the gel, the blocking buffer, and how much antibody was applied to the blot? If I cannot suggest a modification to your protocol, it may be best to replace the antibody with another, which I can offer free-of-charge.
For the GAT2 antibody, ab2896, the precursor is expected to run at 68 kDa. There are several sites on the GAT2 protein that are glycosylated, so it will likely run on the gel to somewhere between 68 and 75 kDa, assuming there are no other post-translation modifications.
Answered on Apr 06 2012