Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor in the gastrointestinal tract. The diagnosis of GIST is based on histology together with a panel of immunohistochemical markers; the most important is KIT (CD117). A total of 434 cases of GISTs were confirmed by histology and immunohistochemistry, and incorporated into tissue microarrays. Validation of histological features as well as the prognostic value of two immunohistochemical biomarkers (p16 and L1) was assessed. High mitotic rate, large tumor size, nuclear atypia, and small bowel primary site were all validated as negative prognostic factors in GISTs. Expression of p16 was significantly correlated with unfavorable prognosis, whereas L1 expression was not.