World-Class Quality | Superior Customer Support | Outstanding Value Tel: 1.858.768.5800 biolegend.com 07-0108-00i Direct-Blot™ HRP Primary Antibodies for Western Blotting When performing a western blot, it is common to use a secondary antibody to detect the proteins of interest. However, when the signal doesn’t need to be amplified, there is no advantage in using a secondary conjugated antibody. Thus, we are happy to introduce our new Horse Radish Peroxidase (HRP) directly conjugated primary antibodies, Direct-Blot™, to help you perform western blots in a breeze. Our HRP directly conjugated antibodies are carefully developed and tested, so sensitivity is not compromised as compared to a traditional western blot using a conjugated secondary antibody. Advantages: • Shorter protocol • Highly sensitive • Excellent stability • Easier to multiplex • Easier to combine with Immunoprecipitation • No potential background from HRP conjugated secondary antibodies To learn more, visit: biolegend.com High sensitivity Western Blot Protocol 15 µg of total protein from either HeLa cells or Jurkat cells treated with PMA (anti- phospho ERK) was resolved by electrophoresis, transferred to nitrocellulose, and probed with different concentrations of HRP directly conjugated antibodies (Direct-Blot™) or purified monoclonal (Pur) anti-GAPDH (clone FF26A/F9 ), anti-Vimentin (clone O91D3), anti-ERK1/2 Phospho (Thr202/Tyr204) (clone 4B11B69) antibodies at a fixed concentration (0.5 µg/mL). For Direct-Blot™ antibodies, proteins were visualized using chemiluminescence detection directly. For purified antibody, proteins were visualized using a goat anti-mouse-IgG secondary antibody conjugated to HRP (Cat. No. 405306) followed by chemiluminescence detection. HeLa cells Jurkat cells