Dataset Information

Purification, identification and profiling of serum amyloid A proteins from sera of advanced-stage cancer patients.

ABSTRACT: Surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-TOF-MS) is a powerful tool for screening potential biomarkers of various pathological conditions. However, low resolution and mass accuracy of SELDI-TOF-MS remain a major obstacle for determination of biological identities of potential protein biomarkers. We report here a refined workflow that combines ZipTip desalting, acetonitrile precipitation, high-performance liquid chromatography (HPLC) separation and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) analysis for the profiling, purification and identification of the targeted serum proteins found by SELDI-TOF-MS. By using this workflow, we purified ten targeted proteins from the sera of patients with various types of advanced stage (stage III-IV) cancers. These proteins were identified as isoforms of the human serum amyloid protein A (SAA) family with or without truncations at their N-terminals. This was confirmed by Western blot analysis. Different SAA expression patterns were observed by MALDI-TOF-MS profiling. SAA has long been reported as a biomarker for various cancer types such as lung cancer, ovarian cancer, and breast cancer. However, in this study we found increased SAA expression in the sera of advanced-stage cancer patients with different cancer types. Our results suggest that maybe SAA should not be used alone as a biomarker for any specific cancer type.

SUBMITTER: Li J

PROVIDER: S-EPMC7105184 | biostudies-literature | 2012 Mar

REPOSITORIES: biostudies-literature

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Purification, identification and profiling of serum amyloid A proteins from sera of advanced-stage cancer patients.

Li Jing J Xie Zhensheng Z Shi Linan L Zhao Zhiqiang Z Hou Junjie J Chen Xiulan X Cui Ziyou Z Xue Peng P Cai Tanxi T Wu Peng P Guo Sutang S Yang Fuquan F

Journal of chromatography. B, Analytical technologies in the biomedical and life sciences 20120116

Surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-TOF-MS) is a powerful tool for screening potential biomarkers of various pathological conditions. However, low resolution and mass accuracy of SELDI-TOF-MS remain a major obstacle for determination of biological identities of potential protein biomarkers. We report here a refined workflow that combines ZipTip desalting, acetonitrile precipitation, high-performance liquid chromatography (HPLC) separation and matr ...[more]

PMID: 22341353

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Project description:BACKGROUND: Schistosomiasis is a chronic disease caused by trematode flatworms of the genus Schistosoma. The disease remains a serious public health problem in endemic countries and affects at least 207 million people worldwide. A definite diagnosis of the disease plays a key role in the control of schistosomiasis. The detection of schistosome circulating antigens (CAs) is an effective approach to discriminate between previous exposure and current infection. Different methods have been investigated for detecting the CAs. However, the components of the schistosome CAs remain unclear. In this study, we analyzed the CAs in sera of patients infected with Schistosoma japonicum. METHODS: The parasites were collected from the infected rabbits for preparing the adult worm antigen (AWA). The hyline hens were immunized subcutaneously with AWA to produce anti-AWA IgY. The IgY was purified by water-dilution and ammonium sulfate precipitation method and identified by ELISA and Western blotting. After purification and characterization, IgY was immobilized onto the resin as a capture antibody. The circulating antigens were immune-precipitated from patients' serum samples by direct immunoprecipitation. The precipitated proteins were separated by one-dimensional electrophoresis and analyzed by LC-MS/MS. RESULTS: Firstly, the IgY against AWA was produced from the eggs of immunized hens by AWA, which gave a titer of 1:12800. The purified IgY was used as the capture antibody to enrich the CAs in sera of S. japonicum infected patients through immunoprecipitation. The CAs were determined by LC-MS/MS. There were four proteins, including protein BUD31 homolog, ribonuclease, SJCHGC06971 protein and SJCHGC04754 protein, which were identified among the CAs. CONCLUSIONS: We developed a novel method based on IgY for identification and profiling CAs in sera of S. japonicum infected patients. Four new CAs were identified and have potential value for further development of an antigen assay.

Dataset Information

Purification, identification and profiling of serum amyloid A proteins from sera of advanced-stage cancer patients.

Publications

Purification, identification and profiling of serum amyloid A proteins from sera of advanced-stage cancer patients.

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