Age-dependent changes in the cerebrospinal fluid proteome by slow off-rate modified aptamer array.

An important precondition for the successful development of diagnostic assays of cerebrospinal fluid (CSF) biomarkers of age-related neurodegenerative diseases is an understanding of the dynamic nature of the CSF proteome during the normal aging process. In this study, a novel proteomic technology was used to quantify hundreds of proteins simultaneously in the CSF from 90 cognitively normal adults 21 to 85 years of age. SomaLogic's highly multiplexed proteomic platform can measure more than 800 proteins simultaneously from small volumes of biological fluids using novel slow off-rate modified aptamer (SOMAmer) protein affinity reagents with sensitivity, specificity, and dynamic ranges that meet or exceed those of enzyme-linked immunosorbent assays. In the first application of this technology to CSF, we detected 248 proteins that possessed signals greater than twofold over background. Several novel correlations between detected protein concentrations and age were discovered that indicate that both inflammation and response to injury in the central nervous system may increase with age. Applying this powerful proteomic approach to CSF provides potential new insight into the aging of the human central nervous system that may have utility in discovering new disease-related changes in the CSF proteome.

From SOMAmer-based biomarker discovery to diagnostic and clinical applications: a SOMAmer-based, streamlined multiplex proteomic assay.

Recently, we reported a SOMAmer-based, highly multiplexed assay for the purpose of biomarker identification. To enable seamless transition from highly multiplexed biomarker discovery assays to a format suitable and convenient for diagnostic and life-science applications, we developed a streamlined, plate-based version of the assay. The plate-based version of the assay is robust, sensitive (sub-picomolar), rapid, can be highly multiplexed (upwards of 60 analytes), and fully automated. We demonstrate that quantification by microarray-based hybridization, Luminex bead-based methods, and qPCR are each compatible with our platform, further expanding the breadth of proteomic applications for a wide user community.

The stability of the circulating human proteome to variations in sample collection and handling procedures measured with an aptamer-based proteomics array.

Blood-based protein biomarkers hold great promise to advance medicine with applications that detect and diagnose diseases and aid in their treatment. We are developing such applications with our proteomics technology that combines high-content with low limits of detection. Biomarker discovery relies heavily on archived blood sample collections. Blood is dynamic and changes with different sampling procedures potentially confounding biomarker studies. In order to better understand the effects of sampling procedures on the circulating proteome, we studied three sample collection variables commonly encountered in archived sample sets. These variables included (1) three different sample tube types, PPT plasma, SST serum, and Red Top serum, (2) the time from venipuncture to centrifugation, and (3) the time from centrifugation to freezing. We profiled 498 proteins for each of 240 samples and compared the results by ANOVA. The results found no significant variation in the measurements for most proteins (approximately 99%) when the two sample processing times tested were 2h or less, regardless of sample tube type. Even at the longest timepoints, 20 h, approximately 82% of the proteins, on average for the three collection tube types, showed no significant change. These results are encouraging for proteomic biomarker discovery.

Aptamer-based multiplexed proteomic technology for biomarker discovery.

BACKGROUND: The interrogation of proteomes ("proteomics") in a highly multiplexed and efficient manner remains a coveted and challenging goal in biology and medicine. METHODOLOGY/PRINCIPAL FINDINGS: We present a new aptamer-based proteomic technology for biomarker discovery capable of simultaneously measuring thousands of proteins from small sample volumes (15 µL of serum or plasma). Our current assay measures 813 proteins with low limits of detection (1 pM median), 7 logs of overall dynamic range (~100 fM-1 µM), and 5% median coefficient of variation. This technology is enabled by a new generation of aptamers that contain chemically modified nucleotides, which greatly expand the physicochemical diversity of the large randomized nucleic acid libraries from which the aptamers are selected. Proteins in complex matrices such as plasma are measured with a process that transforms a signature of protein concentrations into a corresponding signature of DNA aptamer concentrations, which is quantified on a DNA microarray. Our assay takes advantage of the dual nature of aptamers as both folded protein-binding entities with defined shapes and unique nucleotide sequences recognizable by specific hybridization probes. To demonstrate the utility of our proteomics biomarker discovery technology, we applied it to a clinical study of chronic kidney disease (CKD). We identified two well known CKD biomarkers as well as an additional 58 potential CKD biomarkers. These results demonstrate the potential utility of our technology to rapidly discover unique protein signatures characteristic of various disease states. CONCLUSIONS/SIGNIFICANCE: We describe a versatile and powerful tool that allows large-scale comparison of proteome profiles among discrete populations. This unbiased and highly multiplexed search engine will enable the discovery of novel biomarkers in a manner that is unencumbered by our incomplete knowledge of biology, thereby helping to advance the next generation of evidence-based medicine.