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Research

Proteomics Analysis of Human Amniotic Fluid ^*,S

Chan-Kyung J. Cho^,, Shannon J. Shan^,, Elizabeth J. Winsor and Eleftherios P. Diamandis^,,¶,||

From the Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario M5G 1L5, Canada, Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario M5T 3L9, Canada, and ^¶ Department of Clinical Biochemistry, University Health Network and Toronto Medical Laboratories, Toronto, Ontario M5G 2C4, Canada

Amniotic fluid is a dynamic and complex mixture that reflects the physiological status of the developing fetus. In this study, the human amniotic fluid (AF) proteome of a 16–18-week normal pregnancy was profiled and analyzed to investigate the composition and functions of this fluid. Due to the complexity of AF, we utilized three different fractionation strategies to provide greater coverage. Two types of two-dimensional LC/MS/MS as well as an LC-SDS-PAGE-LC-MS/MS platform were used. A total of 16 AF samples between gestational ages of 16 and 18 weeks from women carrying chromosomally normal fetuses were analyzed by one of the three fractionation methods followed by a common reverse phase LC-MS/MS step. Mascot and The Global Proteome Machine engines were used to search the International Protein Index human database for peptide sequence identification. The list of proteins was generated by combining the results of both engines through the PeptideProphet of Scaffold software. All identified proteins were combined to generate the AF proteome comprising 1,026 unique gene matches or 842 non-redundant proteins. This list includes most of the currently used biomarkers for pregnancy-associated pathologic conditions such as preterm delivery, intra-amniotic infection, and chromosomal anomalies of the fetus. The subcellular localization, tissue expression, functions, and networks of the AF proteome were analyzed by various bioinformatic tools. These data will contribute to the better understanding of amniotic fluid function and to the discovery of novel biomarkers for prenatal diagnosis of fetal abnormalities.