title: ProteinSeq: high-performance proteomic analyses by proximity ligation and next generation sequencing.
abstract: Despite intense interest, methods that provide enhanced sensitivity and specificity in parallel measurements of candidate protein biomarkers in numerous samples have been lacking. We present herein a multiplex proximity ligation assay with readout via realtime PCR or DNA sequencing (ProteinSeq). We demonstrate improved sensitivity over conventional sandwich assays for simultaneous analysis of sets of 35 proteins in 5 µl of blood plasma. Importantly, we observe a minimal tendency to increased background with multiplexing, compared to a sandwich assay, suggesting that higher levels of multiplexing are possible. We used ProteinSeq to analyze proteins in plasma samples from cardiovascular disease (CVD) patient cohorts and matched controls. Three proteins, namely P-selectin, Cystatin-B and Kallikrein-6, were identified as putative diagnostic biomarkers for CVD. The latter two have not been previously reported in the literature and their potential roles must be validated in larger patient cohorts. We conclude that ProteinSeq is promising for screening large numbers of proteins and samples while the technology can provide a much-needed platform for validation of diagnostic markers in biobank samples and in clinical use.
authors: Department of Immunology Genetics and Pathology, Uppsala University, Uppsala, Sweden.
authors: Spyros
authors: S
authors: Darmanis
authors: Rachel Yuan
authors: RY
authors: Nong
authors: Johan
authors: J
authors: Vänelid
authors: Agneta
authors: A
authors: Siegbahn
authors: Olle
authors: O
authors: Ericsson
authors: Simon
authors: S
authors: Fredriksson
authors: Christofer
authors: C
authors: Bäcklin
authors: Marta
authors: M
authors: Gut
authors: Simon
authors: S
authors: Heath
authors: Ivo Glynne
authors: IG
authors: Gut
authors: Lars
authors: L
authors: Wallentin
authors: Mats G
authors: MG
authors: Gustafsson
authors: Masood
authors: M
authors: Kamali-Moghaddam
authors: Ulf
authors: U
authors: Landegren
mesh_terms: D015415:Biomarkers / Q000097:blood; D001798:Blood Proteins / Q000032:analysis / Q000235:genetics; D006801:Humans; D007118:Immunoassay / Q000191:economics / Q000379:methods*; D015999:Multivariate Analysis; D040901:Proteomics / Q000191:economics / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000191:economics / Q000379:methods*; D013997:Time Factors
chemical_list: D015415:Biomarkers; D001798:Blood Proteins