Why am I detecting high background with Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127?

Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb #5127 only reacts with native IgG and does not bind to denatured or reduced rabbit IgG heavy chain or light chain. If the antibody used for pull-down in an IP is not fully denatured, it is possible for #5127 to detect IgG fragments, resulting in additional bands. This background may appear as primarily intact antibody around 150 kDa or partially denatured heavy or light chain around 50 kDa or 25 kDa, respectively. 

To resolve additional background around those molecular weights, ensure you are using fresh SDS+DTT sample buffer and heating the samples to 95–100°C for 2-5 minutes prior to running your western blot with #5127 as the secondary.

Last updated: November 21, 2024