Nano‐Biosensors for mRNA‐Based Cell Sorting Using Intracellular Markers at the Early Stage of Cell Reprogramming

Abstract Cell reprogramming and manufacturing have broad applications in tissue regeneration and disease treatment. However, many derived cell types lack unique cell surface markers for protein‐based cell sorting, making it difficult to isolate these cells from mixed populations. Additionally, there is a need to identify and isolate cells of interest at the early stages of cell expansion. To address this challenge, a nucleic acid‐based gold nanorod (NAGNR) fluorescent biosensor was engineered to detect the mRNA expression of intracellular markers for cell sorting. Its application is demonstrated in isolating induced neuronal (iN) cells from dermal fibroblast populations during the early stages of cell reprogramming. Cell sorting based on the mRNA of the neuronal transcriptional factor Ascl1 resulted in an enrichment of iN cells from 3% to 72%, and additional sorting with the transcriptional factor Scn2 further increased iN enrichment. Moreover, NAGNR biosensors can be used in conjunction with protein marker‐based cell sorting. NAGNR‐sorted iN cells show a functional response to electrical stimulation in a co‐culture of iN cells and muscle cells. These findings demonstrate that NAGNR‐based cell sorting offers great potential for cell identification and isolation at an early stage of cell reprogramming and manufacturing.