肝細胞癌(hepatocellular carcinoma，HCC)是種原發性肝癌，同時也是種常見的癌症，尤其是在東南亞以及非洲。腫瘤中有一群具幹細胞特性之癌細胞，有造成腫瘤之能力，比一般癌症細胞更容易產生抗藥性並與腫瘤復發及轉移息息相關，稱之為癌症幹細胞。本實驗主要目的為探討人類肝癌細胞株Hep3B中癌症類幹細胞之特異蛋白質，透過分析蛋白質體之差異性表現，有助於瞭解肝癌進展。
　　運用同重元素相對與絕對定量標記法(isobaric tags for relative and absolute quantitation，iTRAQ®)比較癌症細胞以及不同特性之癌症類幹細胞：可形成球體、少許細胞數即可在小鼠中生長、具抗藥性以及擁有幹細胞表現之特性。從癌症細胞株和癌症類幹細胞株分別萃取一對蛋白質樣品，經還原、烷化、胰蛋白酶水解成胜肽，再標記上iTRAQ®試劑，進一步，將標示上iTRAQ®試劑的胰蛋白酶水解胜肽混合，利用強陽離子交換樹脂(SCX)、等電聚焦分級分離儀(OFFGEL)和鹼性逆相層析(basic reversed-phase chromatography)分餾樣品，接著進入奈米級液相層析電噴灑游離串聯質譜儀(nanoLC ESI tandem MS)偵測。由獲得的特定胜肽，針對每一標記產生之訊號離子質荷比(m/z=114-117)的面積來做相對定量分析，同時胜肽碎片離子(b-及y-型離子)可做為蛋白質鑑定資訊。
　　結合所有分餾樣品，總計約有1,273個蛋白質被 ProteinPilot 軟體鑑定及定量；其中有20和30個蛋白質在癌症細胞及癌症類幹細胞間分別呈現出大於兩倍及小於一半之差異。最後，將這些具顯著性差異的50個蛋白質使用GeneGo軟體進行分析，發現這些差異性表現蛋白質與肝腫瘤、原發性肝癌以及肝病變有所關連，並在代謝、免疫反應以及細胞骨架重組路徑中被發現。在未來，將進行功能性的驗證。

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