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Recombinant Human Adenine Nucleotide Translocase 1.
The following is a quick reference procedure guide for use specifically with ab110322.
1. Solubilize samples in SDS-PAGE sample buffer.
2. Heat sample to 37°C for 30 minutes.
3. Separate proteins on Tris-glycine 10-22% gradient gel 150 V ~ 2 hrs.
4. Soak gel in 50 mL CAPS buffer for 30 minutes.
5. Transfer to 0.45 µm PVDF membrane in CAPS buffer for 2 hrs at 0.15 A CAPS buffer : 10 mM 3-[cyclohexylamino]-1-propane sulfonic acid, pH 11 w/NaOH, 10% methanol ice cold throughout.
6. Block membrane for 3 hrs - overnight in 5% milk/PBS 4°C.
7. Wash blot 3 times for 5 mins PBS/0.05% Tween-20 with agitation. Followed by a final rinse in PBS.
8. Antibody diluted to concentration specified in supplied incubation solution for 2 hrs.
9. Wash blot 3 times in PBS/0.05% Tween-20 for 5 mins.
10. Incubate blot with secondary anti-mouse antibody diluted according to manufacturer’s instructions in 1% milk/PBS for 2 hrs.
11. Wash blot 3 times in PBS/0.05% Tween-20 for 5 mins. Rinse blot in PBS.
12. Develop Blot: For alkaline phosphatase we recommend BCIP/NBT. For horseradish peroxidase we recommend ECL+.
Our Abpromise guarantee covers the use of ab110322 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 1 µg/ml. Predicted molecular weight: 33 kDa.|
|ICC/IF||Use a concentration of 10 µg/ml. (heat-induced antigen-retrieval improves signal)|
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"