A multiplexed assay by self-assembled dual-target responsive DNA hydrogels for efficacy evaluation of immunotherapy

Immunotherapy has revolutionized cancer treatment, yet its efficacy is limited to a specific patient subset. This underscores the critical clinical demand for accessible assays capable of assessing immunotherapy outcomes and enabling timely adjustments to personalized medical care. Here, we present a multiplexed assay based on dual-target responsive DNA hydrogels for the simultaneous detection of soluble programmed death-ligand 1 and lactate dehydrogenase in lung cancer patients. The DNA hydrogel, constructed through a self-assembly strategy, achieves superior performance by reducing background noise by 33.8% and improving the signal-to-noise ratio by 61.5%. By integrating rolling circle amplification, the assay enables ultrasensitive detection at femtomolar levels. When further combining additional clinical biomarkers, the assay demonstrates strong predictive ability for immunotherapy response, achieving an area under the curve value of 0.935 in clinical cohort. Collectively, this blood-based assay offers a versatile and effective approach for advancing biomarker-based evaluations of immunotherapy outcomes.