Urine profiling by SELDI-TOF/MS: Monitoring of the critical steps in sample collection, handling and analysis
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- 作者：Massimo Papale ; Maria Carmela Pedicillo ; Bradley J. Thatcher ; Salvatore Di Paolo ; Lorenzo Lo Muzio ; Pantaleo Bufo ; Maria Teresa Rocchetti ; Marta Centra ; Elena Ranieri ; Loreto Gesualdo
- 关键词：Urine ; Sample collection ; SELDI-TOF/MS ; Standardization
- 刊名：Journal of Chromatography B, Analytical Technologies in the Biomedical and Life Sciences
- 出版年：2007
- 出版时间：1 September 2007
- 年：2007
- 卷：856
- 期：1-2
- 页码：205-213
- 全文大小：557 K
文摘
The topic of this study is the impact of several pre-analytical and analytical variables on proteomic profiling of human urine by surface enhanced laser desorption/ionization time of flight–mass spectrometry (SELDI-TOF–MS) in healthy subjects. Urine storage at room temperature caused a progressive degradation of proteins, which was prevented by the addition of protease inhibitors only up to 2 h from the collection. The timing of collection over the day had only a minor impact on protein profile, although influencing the intensity of peaks. Repeated freeze/thaw cycles (up to five) did not affect either the number or the intensity of the peaks. A comparison of the protein profile from eight different healthy individuals showed fairly consistent inter-subject similarities, along with between-subject differences, which were markedly dependent on the sex and the type of ProteinChip® array used. The addition of a variety of denaturing agents improved the quality of the spectra with all the chips tested (CM10, Q10 and H50), but not with the copper-coated IMAC-30 chip. Finally, SPA matrix allowed to achieve a better performance of SELDI-TOF/MS spectrum, as compared with CHCA, regardless of the ProteinChip® array used and even in the low m/z range (2500–10,000). In conclusion, we suggest that a careful choice of a number of pre-analytical and analytical conditions is required to accomplish and define a unifying protocol for the analysis of human urine by SELDI-TOF/MS, in physiological and in pathological states.