Peptide Aptamer-Enabled Nanoplasmonic Digital Immunoassay for Ultrasensitive Cytokine Sensing in Early Inflammation and Immune Modulation.

Early and precise detection of cytokine release is essential for mitigating cytokine release syndrome, a life-threatening complication of CAR T-cell therapy. However, existing immunoassays fall short in sensitivity, dynamic range, and temporal resolution, limiting their ability to capture the subtle, transient cytokine fluctuations that mark the earliest phase of immune activation. To address this unmet need, we developed an ultrasensitive nanoplasmonic digital immunoassay that integrates rationally engineered antibody-derived peptide aptamers (ADPAs), plasmonic gold nanospheres for digital dark-field imaging, and convolutional neural network signal quantification. This platform enables ultrasensitive cytokine detection down to tens of fg/mL, with quantitative coverage across the tested concentration range spanning approximately six orders of magnitude using only microliter-scale sample volumes. We applied this technology to a physiologically relevant in vitro model of CAR T-cell-induced cytokine release, enabling high-frequency cytokine monitoring at early time points previously inaccessible to conventional assays. More broadly, this platform enables continuous, ultra-sensitive monitoring of cytokine dynamics at the onset of inflammation, opening new opportunities for early diagnosis and precision-guided intervention in a wide range of immune-mediated conditions.