Two-dimensional Northwestern blotting

Two dimensional (2D) Northwestern blotting complements gel-shift experiments in the identification of specific protein-nucleic acid interactions. The procedures described below, which have been adapted from the protocol described by Schenkel et al. (1988), have been valuable in our laboratory for the identification of several hnRNP proteins (our unpublished observations). Here, we demonstrate the technique on 2D gel blots of transformed human amnion (AMA) cell proteins (Fig. 1) using poly(rC), a homopolymer that interacts with hnRNPs K and L (Matunis et al., 1992 Dejgaard et al., 1994), as well as poly(rU), which interact with hnRNPs C and M (Swanson and Dreyfuss, 1988 Dreyfuss et al., 1993). The technique works equally well, however, when sequence-specific deoxyoligonucleotides are used as probes. 

---