Quantitative proteomic analysis of formalin-fixed and paraffin-embedded nasopharyngeal carcinoma using iTRAQ labeling, two-dimensional liquid chromatography, and tandem mass spectrometry.
Formalin-fixed, paraffin-embedded (FFPE) tissue specimens represent a potentially valuable resource for protein biomarker investigations. In this study, proteins were extracted by a heat-induced antigen retrieval technique combined with a retrieval solution containing 2% SDS from FFPE tissues of normal nasopharyngeal epithelial tissues (NNET) and three histological types of nasopharyngeal carcinoma (NPC) with diverse differentiation degrees. Then two-dimensional liquid chromatography-tandem mass spectrometry coupled with isobaric tags for relative and absolute quantification (iTRAQ) labeling was employed to quantitatively identify the differentially expressed proteins among the types of NPC FFPE tissues. Our study resulted in the identification of 730 unique proteins, the distributions of subcellular localizations and molecular functions of which were similar to those of the proteomic database of human NPC and NNET that we had set up based on the frozen tissues. Additionally, the relative expression levels of cathepsin D, keratin8, SFN, and stathmin1 identified and quantified in this report were consistent with the immunohistochemistry results acquired in our previous study. In conclusion, we have developed an effective approach to identifying protein changes in FFPE NPC tissues utilizing iTRAQ technology in conjunction with an economical and easily accessible sample preparation method.
Xiao
Zhefeng
Z
Key Laboratory of Cancer Proteomics of Chinese Ministry of Health, Xiangya Hospital, Central South University, 87# Xiangya Road, Changsha 410008, Hunan, China.
Li
Guoqing
G
Chen
Yongheng
Y
Li
Maoyu
M
Peng
Fang
F
Li
Cui
C
Li
Feng
F
Yu
Yanhui
Y
Ouyang
Yongmei
Y
Xiao
Zhiqiang
Z
Chen
Zhuchu
Z
D048948:14-3-3 Proteins; D000595:Amino Acid Sequence; D014408:Biomarkers, Tumor / Q000378:metabolism; D002402:Cathepsin D / Q000378:metabolism; D002454:Cell Differentiation; D002853:Chromatography, Liquid / Q000379:methods; D005092:Exonucleases / Q000378:metabolism; D005095:Exoribonucleases; D005557:Formaldehyde; D006801:Humans; D008969:Molecular Sequence Data; D009303:Nasopharyngeal Neoplasms / Q000378:metabolism* / Q000473:pathology; D009363:Neoplasm Proteins / Q000302:isolation & purification / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D050777:Stathmin / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism / Q000473:pathology; D053719:Tandem Mass Spectrometry / Q000379:methods
D048948:14-3-3 Proteins; D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D010446:Peptide Fragments; C493995:STMN1 protein, human; D050777:Stathmin; D005557:Formaldehyde; D005092:Exonucleases; D005095:Exoribonucleases; C413442:SFN protein, human; C533922:CTSD protein, human; D002402:Cathepsin D