Tissue microarray(TMA) construction and immunohistochemistry(IHC) staining

The TMA used for this study includes unselected, non-consecutive, primary, and sporadic colorectal cancers enrolled between January 2007 and November 2009 in FUSCC. Construction of this TMA has been previously described in detail [14]. Every section was scored independently by two pathologists utilizing a semiquantitative scoring system [15, 16]. The staining intensity was scored as 0 (negative), 1 (weak), 2 (medium) or 3 (strong). Extent of staining was scored as 0 (< 5%), 1(5–25%), 2 (26–50%), 3 (51–75%) and 4 (> 75%) according to the percentages of the positive staining areas in relation to the whole carcinoma area. We multiplied the percentage score by the staining intensity score to generate the immunoreactivity score (IRS). High expression of OGN was defined as detectable immunoreactions in cytoplasm and stoma with IRS > 4.