Novel Homo-SELEX for selecting universal aptamer for cross-species kidney injury biomarker KIM-1 in muti-scenario situ imaging.

Kidney injury molecule 1 (KIM-1) has emerged as a critical biomarker for renal injury, playing a pivotal role in disease diagnosis and therapeutic monitoring. Despite its clinical importance, the development of high-affinity molecular probes for KIM-1 remains challenging. Aptamers are valuable in biorecognition because of their easy synthesis, low toxicity and immunogenicity. The traditional Cell-SELEX suffers from poor cross-species compatibility, hindering its utility for KIM-1 detection. To overcome these challenges, we developed Homo-SELEX, an innovative cell-free SELEX strategy designed to screen aptamers against membrane proteins across multiple species. After 15 rounds of selection, universal aptamer S7 exhibited the highest affinity with different species of KIM-1. Aptamer S4 has specificity in identifying KIM-1 from homo sapiens and murine. The kidney tissue imaging demonstrated that S7 exhibited a better affinity than the S4. Furthermore, imaging of human clinical renal membranitis paraffin tissues, chronic kidney disease paraffin tissues of rats and acute nephrotoxic frozen tissues of mice, aptamer S7 exhibited a specific recognition of KIM-1. We also demonstrated the feasibility of S7 aptamer targeting KIM-1 in lung cancer A549 and liver injury tissues of rat and mouse, while demonstrating no binding to normal HK2 and tissue. Our works establish Homo-SELEX as a breakthrough strategy for homologous protein screening, with S7 serving as a robust, species-agnostic molecular probe for KIM-1 detection. This advancement holds significant promise for non-invasive diagnostics and targeted therapies for KIM-1-associated pathologies.