ORIGINAL PAPER 14-3-3 Protein beta/alpha as a urinary biomarker for renal cell carcinoma: proteomic analysis of cyst fluid Satoru Minamida & Masatsugu Iwamura & Yoshio Kodera & Yusuke Kawashima & Kenichi Tabata & Kazumasa Matsumoto & Tetsuo Fujita & Takefumi Satoh & Tadakazu Maeda & Shiro Baba Received: 28 January 2011 /Revised: 6 April 2011 /Accepted: 22 April 2011 /Published online: 8 May 2011 # Springer-Verlag 2011 Abstract Although various samples, including tissue, cells, serum, and urine, from patients with renal cell carcinoma (RCC) have been analyzed, biomarkers with diagnostic value have yet to be identified. We used a proteomics approach to analyze cyst fluid in cases of cyst-associated RCC to identify accessible and abundant proteins that are overexpressed and/or secreted by RCC cells. Proteins in the cyst fluid were separated by reverse-phase high- performance liquid chromatography and agarose two- dimensional gel electrophoresis and were identified by tandem mass spectrometry. We conducted a National Center for Biotechnology Information search and a MEDLINE search to predict the function of these identified proteins and to select a tumor-marker candidate protein. Our search resulted in the identification and selection of the differen- tially regulated protein known as 14-3-3 protein beta/alpha, which was overexpressed in cyst fluid from cyst-associated RCC but has not been previously associated with RCC. We then measured its incidence through Western blotting of various normal and RCC samples (serum, urine, tissue, and cyst fluid). The expression levels of 14-3-3 protein beta/ alpha were higher in urine samples from patients with RCC than in samples from healthy volunteers. Receiver operat- ing characteristic (ROC) curve analyses were performed to assess this potential biomarker; these data (area under the ROC curve value was 0.8813) indicate a high degree of accuracy for this screening method. 14-3-3 Protein beta/ alpha may be a diagnostically useful biomarker for early diagnosis of RCC. Keywords Renal cancer . Proteome . Marker . 14-3-3 Protein . Cyst fluid Introduction Because renal cell carcinoma (RCC) can be resistant to chemotherapy and radiotherapy, nephrectomy is often the most effective treatment option [1]. Although nephrectomy can be expected to cure patients without metastatic involvement, 15–25% of RCC patients already have metastatic RCC at the time of diagnosis [2]. Once metastatic disease develops, the prognosis for long-term survival is poor: The 5-year survival rate for those with metastasis is less than 10%. Because RCC often presents with few signs, symptoms, or laboratory abnormalities, it is often diagnosed in later stages of the disease. An increasing percentage of RCC tumors are detected through imaging performed because of symptoms unrelated to RCC. At the same time, the incidence of RCC and the rate of mortality due to RCC are both increasing [3, 4], primarily due to the lack of a useful biomarker that can detect early stage RCC. Cancer proteomics is expected to be useful in the search for new RCC biomarkers in serum and urine samples, which could eventually serve as a means of early cancer diagnosis. Proteomics analysis has been used to identify potential biomarkers for RCC, and various putative candi- Electronic supplementary material The online version of this article (doi:10.1007/s00216-011-5057-5) contains supplementary material, which is available to authorized users. S. Minamida : M. Iwamura (*) : K. Tabata : K. Matsumoto : T. Fujita : T. Satoh : S. Baba Department of Urology, Kitasato University School of Medicine, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa 252–0374, Japan e-mail: dm08025e@st.kitasato-u.ac.jp Y. Kodera : Y. Kawashima : T. Maeda Department of Physics, Kitasato University School of Science, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa 252–0374, Japan Anal Bioanal Chem (2011) 401:245–252 DOI 10.1007/s00216-011-5057-5