2D MOF Nanosensor‐Integrated Digital Droplet Microfluidic Flow Cytometry for In Situ Detection of Multiple miRNAs in Single CTC Cells

Abstract

Current circulating tumor cells (CTCs) detection strategies based on surface epithelial markers suffer from low specificity in distinguishing between CTCs and epithelial cells in hematopoietic cell population. Tumor‐associated miRNAs within CTCs are emerging as new biomarkers due to their high correlation with tumor development and progress. However, in‐situ simultaneous analysis of multiple miRNAs in single CTC cell is still challenging. To overcome this limitation, a digital droplet microfluidic flow cytometry based on biofunctionalized 2D metal‐organic framework nanosensor (Nano‐DMFC) is developed for in situ detection of dual miRNAs simultaneously in single living breast cancer cells. Here, 2D MOF‐based fluorescent resonance energy transfer (FRET) nanosensors are established by conjugating dual‐color fluorescence dye‐labeled DNA probes on MOF nanosheet surface. In the Nano‐DMFC, 2D MOF‐based nanoprobes are precisely microinjected into each single‐cell encapsulated droplets to achieve dual miRNA characterization in single cancer cell. This Nano‐DMFC platform successfully detects dual miRNAs at single‐cell resolution in 10 mixed positive MCF‐7 cells out of 10 000 negative epithelial cells in serum biomimic samples. Moreover, this Nano‐DMFC platform shows good reproductivity in the recovery experiment of spiked blood samples, which demonstrate the high potential for CTC‐based cancer early diagnosis and prognosis.

Circulating miR-200 Family and CTCs in Metastatic Breast Cancer before, during, and after a New Line of Systemic Treatment

“…Sensitive readout approaches include SERS (surface-enhanced Raman scattering), SPR (surface plasmon resonance), and electrochemical labeling (e.g., CRISPR-based biosensing platforms) [ 53 ]. Excitingly, miRNAs could recently be measured directly in CTCs in vivo, promising great tumor specific diagnostic, prognostic, and therapeutic potential [ 54 , 55 ]. However, RT-qPCR is undisputedly considered the gold standard for miRNA detection, as it offers attomolar sensitivity and specificity for single cells and is also cost-effective [ 53 ].…”

Advances in Microfluidics: Technical Innovations and Applications in Diagnostics and Therapeutics

“…They demonstrated multiplex detection of 5 representative miRNA biomarkers for breast cancer diagnosis. Similarly, Chen et al used a 2D metal–organic framework nanosensor and FRET to detect microRNAs but in single circulating tumor cells in droplets . Single-cell droplets were generated, injected with the nanosensors, and passed through the fluorescence detector in a flow cytometric analysis.…”

“…Similarly, Chen et al used a 2D metal−organic framework nanosensor and FRET to detect microRNAs but in single circulating tumor cells in droplets. 104 Single-cell droplets were generated, injected with the nanosensors, and passed through the fluorescence detector in a flow cytometric analysis. Ren et al used an electrokinetic sensor based on ion-exchange membranes to detect miRNA for myocardial infarction with a picomolar limit of detection.…”

Developments in FRET- and BRET-Based Biosensors

“…It is conducive to high-throughput detection. Microfluidic has also been applied to RET signal detection, especially in the development of portable detection devices to obtain rapid results using fewer samples [ 65 , 102 , 103 , 104 ]. The use of smartphones to detect RET signals is also a future direction.…”