Development of a heat-mediated protein blotting method

Western blotting is a significant tool employed for the detection of cell proteins. High-molecular-weight proteins have proven a challenge to detect by western blotting, but proteins even of 100 KDa can still present difficulties in detection. This work reports the development of a heat transfer method that is suitable for both low- and high-molecular-weight proteins. The procedure involves the use of a constant temperature at 78 °C in a dedicated heat transfer module. Through the use of this protocol the neuronal adaptor protein X11α (120 KDa), which prior to this methodology was undetectable endogenously in the neuroblastoma cell line (N2a), was successfully detected in the N2a cell line. The procedure provides a reproducible protocol that can be adapted for other high-molecular-weight proteins, and it provides the advantage that low-molecular-weight proteins are not sacrificed by the methodology.