Evaluation and refinement of sample preparation methods for extracellular matrix proteome coverage
The extracellular matrix is a key component of tissues, yet it is under-represented in proteomic datasets. Identification and evaluation of proteins in the extracellular matrix (ECM) has proved challenging due to the insolubility of many ECM proteins in traditional protein extraction buffers. Here we separate the decellularization and ECM extraction steps of several prominent methods for evaluation under real-world conditions. The results are used to optimize a two-fraction ECM extraction method. Approximately one dozen additional parameters are tested and recommendations for analysis based on overall ECM coverage or specific ECM classes are given. Compared to a standard in-solution digest, the optimized method yielded a 4-fold improvement in unique ECM peptide identifications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/391946v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@7e356aorg.highwire.dtl.DTLVardef@1e4f196org.highwire.dtl.DTLVardef@1b0e10forg.highwire.dtl.DTLVardef@187e040_HPS_FORMAT_FIGEXP M_FIG C_FIG