Development of rapid diagnostic kit against sepsis: Barcode-based semi-quantitative measurement of C-reactive protein...

This study reports the development of a point-of-care (POC) detection platform integrating lateral flow immunoassays (LFIAs) and paper-based enzymatic sensors for the semi-quantitative measurement of C-reactive protein (CRP) and lactate, two key biomarkers strongly associated with inflammatory conditions and sepsis. This approach relies on a visual barcode-like gradient readout that enables instrument-free analysis for rapid clinical evaluation. A lateral flow assay employing gold nanoparticles (AuNPs) conjugated to anti-CRP antibodies was designed for CRP detection, while lactate levels were assessed using multi-dilution lactate dehydrogenase (LDH)-embedded paper sensors coupled to a redox dye system. Characterization of AuNP-antibody conjugates by UV-Vis spectroscopy, salt-induced aggregation, and electron microscopy confirmed stable surface functionalization. The LFIA test strips provided a three-line gradient response, enabling semi-quantitative interpretation of CRP levels across clinically relevant thresholds, demonstrating a linear response between 0.2 and 12.5 mg/L (R² = 0.95) with a visual cut-off of 10 mg/L in clinical samples. The lactate paper sensors, employing 2-fold, 5-fold, and 10-fold dilution strategies, generated distinguishable color gradients for various concentrations within a quantitative range of 0.39-9 mM with a coefficient of determination greater than 0.95. For visual interpretation, quantitative image analysis based on pixel intensity was employed to improve the accuracy of paper sensor. The results from the LFIA and paper-based sensors showed a strong correlation with reference laboratory methods, specifically ELISA for CRP and the spectroscopic method for lactate, respectively. Furthermore, stability studies indicated that the paper-based sensors retain over 90% of their initial activity for 16 days at 4 °C, while the LFIA strips remain stable for up to one year at room temperature. The combined biosensor platform provided a gradient-based readout for both biomarkers, enabling rapid, semi-quantitative discrimination across clinically relevant concentration ranges without the need for specialized instrumentation. Overall, this work demonstrates the development of a rapid diagnostic kit capable of detecting blood lactate and CRP levels within a short time and with clinically relevant accuracy.