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A simplified perchloric acid workflow with neutralization (PCA-N) for democratizing deep plasma proteomics at population scale - Submission04

ABSTRACT: This project presents PCA-N, a novel plasma proteomics workflow that uses perchloric acid with neutralization to dramatically improve sample preparation efficiency. By eliminating solid-phase extraction and enabling 384-well plate compatibility, PCA-N reduces sample volume requirements to just 5 μL, cuts costs to under $1 per sample, and allows processing of up to 13,000 samples daily. Validated on over 34,000 samples across 311 days, this method doubles proteome depth compared to conventional approaches while maintaining robust measurement precision, making large-scale plasma proteomics studies feasible for biomarker discovery and precision medicine.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Plasma

SUBMITTER: Mario Oroshi

LAB HEAD: Prof. Dr. Matthias Mann

PROVIDER: PXD064487 | Pride | 2025-09-30

REPOSITORIES: Pride

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Project description:Osteoporosis is the consequence of altered bone metabolism resulting in the systemic reduction of bone strength and increased risk of fragility fractures. MicroRNAs (miRNAs) regulate gene expression on a post-transcriptional level are known to take part in the control of bone formation and bone resorption. Recently, targeted secretion of miRNAs from cells originating from various tissues has been described, which allows for their minimal-invasive detection in serum/plasma and use as biomarkers for presence and progression of pathological conditions. One pilot study has reported circulating miRNAs in serum and tissue of fracture patients. However, further studies are required to explore whether a dysbalance in bone homeostasis of fracture patients can reliably be reflected by specific circulating miRNAs, and whether these miRNAs might serve as drugable targets. Here, we report results from a comprehensive multiplex study of 175 miRNAs in serum samples obtained from 7 patients with osteoporotic fractures at the femoral neck, and 7 age-matched controls. Following elaborate quality control statistical analysis of this exploratory dataset identified 9 microRNAs with altered serum levels in response to fracture (adjusted p-value < 0.1). Of these, hsa-miR-10a/b gave excellent discrimination of both groups (AUC = 1.0), and clustering of samples based on the top10 miRNAs confirmed the high discriminatory power of circulating microRNAs for osteoporotic fractures. In the next step 3 miRNAs with unknown roles in osteogenic differentiation and 4 miRNA from a previous study were tested for their effects on osteogenic differentiation. Of these, 3 miRNAs showed robust effects on osteogenic differentiation. Overall, these data provide important insights into changes in serum miRNA in post-traumatic patients. Future studies will show, whether this knowledge can be used to improve current diagnostic methodologies to predict fracture risk and design novel treatment strategies for osteoporosis patients. Two groups with n=7 per group; one groups represents cases with osteoporotic fractures, the control group is age-matched without fractures

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A simplified perchloric acid workflow with neutralization (PCA-N) for democratizing deep plasma proteomics at population scale - Submission04

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