M001:Micro-scale immunoblotting for small sample protein Analysis

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Authors: Sara Saedinia, Liang (Lily) Wu, Kent Nastiuk, John Krolewski, Mark Bachman

Microscale immunoblotting for small sample protein analysis


Western blot, electrophoretic separation followed by immunoblotting, is a well understood and popular technique for analyzing and quantifying protein composition in a complex mixture. This technique can potentially be useful for identifying the protein expression of cancer cells in resected tumor samples, thus providing important information for applications in personalized medicine for cancer treatment. However, conventional embodiments of this technique typically require large quantities of proteins, making it unsuitable for anayzing small samples of tissue containing tumor cells of interest, which often results in proteins yields measured in nanograms.

We are developing a micro-western blot system that can work on a very small quantity of protein in solution, making it suitable for use with small samples such as cancer cells separated from biopsy tissue samples (such as by laser capture microdissection). The device employs microfluidic gels and electrophoretic transport to pre-concentrate the proteins in a stacking region, then performs protein separation in a long microgel. The small gel dimensions ensure that proteins remain in high concentration during separation, even though the total quantity of the proteins is small. After separation, microfluidic side channels pull the proteins into hydrophobic capture regions ("micro-blotting"), where immunostaining is subsequently performed. This device is thus capable of running a small volume sample through a complete western blot analysis on an integrated "chip". The small integrated device may be driven by external electronics, and so may be completely automated, making it suitable for use by non-specialists, for example, in the field or at the point of care.