Western Blot - Goat Anti-Rabbit IgG H&L Antibody (HRP) (A17345)
Western blot analysis of extracts of HeLa cells, using Goat Anti-Rabbit IgG H&L Antibody (HRP) (A17345) at 1:4000-1:10,000 dilution. Lysates/proteins were present at 25µg per lane. The blocking buffer used was 3% non-fat dry milk in TBST. Detection was with a ECL Basic Kit. Exposure time: 3s.
Experimental Conditions: Proteins were resolved by SDS–PAGE on a 12.5% polyacrylamide gel run at 200 V for 55 minutes, with 10 µl of undiluted protein sample mixed 1:1 with loading dye prior to electrophoresis. Separated proteins were transferred onto a nitrocellulose membrane using the iBlot 2 dry blotting system at 20 V for 5 minutes, in accordance with the manufacturer’s instructions. Transfer efficiency was verified by Ponceau S staining. Membrane blocking and antibody incubations were performed using the iBind system with overnight incubations. Immunoreactive bands were visualised using Immobilon Forte Western HRP substrate and imaged on an iBright imaging system.
Dilution: 1:1000
Results Summary: Clear detection of a poorly expressed membrane protein in B. thetaiotaomicron with strong signal and low background.
Comments: Excellent sensitivity and reliability for detection of low-abundance bacterial membrane proteins under iBind conditions.
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