Research | Children’s Health Prioritization of Biomarker Targets in Human Umbilical Cord Blood: Identification of Proteins in Infant Blood Serving as Validated Biomarkers in Adults Nicole Hansmeier,1 Tzu-Chiao Chao,1,2 Lynn R. Goldman,3 Frank R. Witter,4 and Rolf U. Halden1,5,6 1Swette Center for Environmental Biotechnology, Biodesign Institute, Arizona State University, Tempe, Arizona, USA; 2Department of Chemistry and Biochemistry, Arizona State University, Tempe, Arizona, USA; 3School of Public Health and Health Services, George Washington University, Washington, District of Columbia, USA; 4Department of Gynecology and Obstetrics, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; 5Department of Environmental Health Sciences, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland, USA; 6Center for Health Information and Research, Arizona State University, Tempe, Arizona, USA Background : Early diagnosis represents one of the best lines of defense in the fight against a wide array of human diseases. Umbilical cord blood (UCB) is one of the first easily available diagnostic biofluids and can inform about the health status of newborns. However, compared with adult blood, its diagnostic potential remains largely untapped. o Bjectivs : Our goal was to accelerate biomarker research on UCB by exploring its detectable pro- tein content and providing a priority list of potential biomarkers based on known proteins involved in disease pathways. Methods : We explored cord blood serum proteins by profiling a UCB pool of 12 neonates with different backgrounds using a combination of isoelectric focusing and liquid chromatography cou- pled with matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) and by comparing results with information contained in metabolic and disease databases available for adult blood. r sulte : A total of 1,210 UCB proteins were identified with a protein-level false discovery rate of ~ 5% as estimated by naïve target-decoy and MAYU approaches, signifying a 6-fold increase in the number of UCB proteins described to date. Identified proteins correspond to 138 different meta- bolic and disease pathways and provide a platform of mechanistically linked biomarker candidates for tracking disruptions in cellular processes. Moreover, among the identified proteins, 38 were found to be approved biomarkers for adult blood. c onclusi : hTe results of this study advance current knowledge of the human cord blood serum proteome. They showcase the potential of UCB as a diagnostic medium for assessing infant health by detection and identification of candidate biomarkers for known disease pathways using a global, non- targeted approach. hTese biomarkers may inform about mechanisms of exposure–disease relation - ships. Furthermore, biomarkers approved by the U.S. Food and Drug Administration for screening in adult blood were detected in UCB and represent high-priority targets for immediate validation. k ey words : body fluid, diagnostics, disease, LC-MALDI-MS, pathways, proteomics. Environ Health Perspect 120:764–769 (2012). http://dx.doi.org/10.1289/ehp.1104190 [Online 27 January 2012] Many diseases are of early-life origin. Early diagnosis of diseases, toxic exposures, effects, and susceptibilities in the still-developing body of infants will be required to develop successful intervention and treatment strate- gies to battle diseases. It is well documented that exposures to environmental chemical contaminants, including cigarette smoke con- stituents, for example, have adverse effects on fetal development and result in unfa- vorable health trajectories for affected chil- dren (Apelberg et al. 2007a; Buczyńska and Tarkowski 2005; Mattison 2010; Miranda et al. 2009; Wigle et al. 2007). Long-term outcomes such as diabetes, obesity, and chronic heart and kidney diseases have all been hypothesized or postulated to have their basis in fetal and childhood exposure (Barker et al. 2002) and show an increased prevalence in children and newborns (Bloomgarden 2004; Ferrara 2007). Therefore, the develop- ment of early diagnostics as predictors for child health is of paramount importance to enable early intervention. 764 Umbilical cord blood (UCB) is a very attractive biological specimen, because rela- tively high volumes (up to tens of milliliters) of this biofluid are available for sampling without posing an added risk and burden to the newborn or its mother in the process. In addition, it has the potential to inform about existing or potential future adverse effects. UCB is already in use for prediag- nosis and treatment of immune deficiencies (Notarangelo 2010). However, its primary use at this time is for bone marrow transplan- tation (Buchheiser et al. 2009). Mass spectrometry (MS)–based proteom- ics is a powerful technology, allowing for the identification and quantification of hundreds of proteins in parallel from a single sample without necessitating prior selection or exclu- sion of potential analytical targets. It has been employed successfully for a diverse range of organisms, tissues, and biofluids (Aebersold and Mann 2003; Ahrens et al. 2010a, 2010b; Beck et al. 2011; Domon and Aebersold 2010; Nilsson et al. 2010) and offers an ideal platform for fast identification of new protein markers for diseases or adverse expo- sure (Lemos et al. 2010; Liumbruno et al. 2010). For adult blood, there is a long tradi- tion of analyzing serum proteins using two- dimensional gel electrophoresis (Hughes et al. 1992) or shotgun proteomics (Omenn et al. 2005; Pieper et al. 2003; Richter et al. 1999; Schenk et al. 2008; States et al. 2006), with continuous analytical improvements (Bell et al. 2009; Gaso-Sokac and Josic 2010; Rai et al. 2005). In contrast to adult blood, much less information is available for the cord blood proteome. Recently, two short overviews on cord blood proteomics were published describing 207 and 837 different proteins, respectively (Colquhoun et al. 2009; Song et al. 2009). Unfortunately, the latter report provided information on only 61 of the 837 proteins. The identifiers of the 776 remaining proteins are not published. More detailed investigations of the UCB proteome are needed to accelerate the pace of discovery and expand the spectrum of infant health diagnostics. In addition, the sampling of UCB for proteome studies has to be suc- cessful within the limitations of clinical real- ity and is therefore much more challenging than the analysis of adult blood. The aim of this study was to provide insights into several important aspects necessary to use the UBC as an effective source for protein-based biomark- ers. As a first priority, we aimed to expand the knowledge on the detectable proteome in UCB, which can be obtained from a limited starting volume. This is especially important Address correspondence to R.U. Halden, Swette Center for Environmental Biotechnology, The Biodesign Institute at Arizona State University, PO Box 875701 Tempe, AZ 85287 USA. Telephone: (480) 727-0893. Fax: (480) 727-0889. E-mail: halden@asu.edu Supplemental Material is available online (http:// dx.doi.org/10.1289/ehp.1104190). This project was supported in part by research grant R01ES015445 and its supplements awarded by the National Institute of Environmental Health Sciences (NIEHS). The content of the article is solely the responsibility of the authors and does not necessarily represent the official views of the NIEHS or the National Institutes of Health. The authors declare they have actual or potential competing financial interests. Received 8 July 2011; accepted 27 January 2012. ovlume 120 | number 5 | May 2012 • Environmental Health Perspectives