Engineering Quantum Dots with Different Emission Wavelengths and Specific Fluorescence Lifetimes for Spectrally and Temporally Multiplexed Imaging of Cells

In this work, a proof-of-concept study was performed to examine the potential of spectrally and temporally multiplexed imaging of cells by using quantum dots (QDs). The CdSe and ZAIS QDs with different emission wavelengths and well-separated fluorescence lifetimes were prepared to provide 2-dimensional information. After incubation with cells, the same type of QDs with different emission wavelengths were distinguishable in spectral imaging while different types of QDs with similar emission wavelengths but well-separated fluorescence lifetimes were resolvable in fluorescence lifetime imaging. For cells co-stained with dye and different types of QDs, the fluorescence lifetime imaging microscopy (FLIM) images showed spatially separated patterns that can be split into channel images by using the software-based time gates. Overall, the results demonstrate the feasibility of combining the 2-dimensional encoded QDs for spectrally and temporally multiplexed imaging. This method can be extended to other QDs and organic dyes to maximize the number of measurable species in multiplexed imaging and sensing applications.