In-situ quantification of lipids in live cells through imaging approaches.

Lipids are important molecules that are widely distributed within the cell, and they play a crucial role in several biological processes such as cell membrane formation, signaling, cell motility and division. Monitoring the spatiotemporal dynamics of cellular lipids in real-time and quantifying their concentrations in situ is crucial since the local concentration of lipids initiates various signaling pathways that regulate cellular processes. In this review, we first introduced the historical background of lipid quantification methods. We then delve into the current state of the art of in situ lipid quantification, including the establishment and utility of fluorescence imaging techniques based on sensors of lipid-binding domains labeled with organic dyes or fluorescent proteins, and Raman and magnetic resonance imaging (MRI) techniques that do not require lipid labeling. Next, we highlighted the biological applications of live-cell lipid quantification techniques in the study of in situ lipid distribution, lipid transformation, and lipid-mediated signaling pathways. Finally, we discussed the technical challenges and prospects for the development of lipid quantification in live cells, with the aim of promoting the development of in situ lipid quantification in live cells, which may have a profound impact on the biological and medical fields.

In Situ Quantification of Lipids in Live Cells by Using Lipid-Binding Domain-Based Biosensors.

Lipid molecules contribute to a large extent to the regulation of cellular signaling, as cellular signals are generated primarily through the selective interaction of various cellular proteins with lipids in the plasma membrane. Hence the location, concentration, and duration of lipids on the cell membrane are critical for the selection of proteins and the initiation of signaling. To monitor the concentration and location of lipid molecules on the cell membrane, researchers have developed a variety of lipid biosensors that allow quantitative in situ visualization of lipid molecules in living cells based on lipid-binding domains with high specificity, sensitivity, and biocompatibility, providing a powerful tool for the study of cellular signaling mechanisms involving lipid molecules. In this review, we first introduced the emergence of lipid-binding domains and then focused on the practical considerations on how to implement the lipid sensor, including probe selection, modification, characterization, and imaging techniques. We then described experimental observables and the relevant physicochemical parameters in the context of single-molecule studies in cells. Finally, we presented our views on the future development of lipid sensors and methods for lipid quantification.

Rapid detection and subtyping of multiple influenza viruses on a microfluidic chip integrated with controllable micro-magnetic field.

Influenza viruses have threatened animals and public health systems continuously. Moreover, there are many subtypes of influenza viruses, which have brought great difficulties to the classification of influenza viruses during any influenza outbreak. So it is crucial to develop a rapid and accurate method for detecting and subtyping influenza viruses. In this work, we reported a rapid method for simultaneously detecting and subtyping multiple influenza viruses (H1N1, H3N2 and H9N2) based on nucleic acid hybridization on a microfluidic chip integrated with controllable micro-magnetic field. H1N1, H3N2 and H9N2 could be simultaneously detected in 80min with detection limits about 0.21nM, 0.16nM, 0.12nM in order. Moreover, the sample and reagent consumption was as low as only 3µL. The results indicated that this approach possessed fast analysis and high specificity. Therefore, it is expected to be used to simultaneously subtype and detect multiple targets, and may provide a powerful technique platform for the rapid detection and subtyping analysis of influenza viruses.

A simple point-of-care microfluidic immunomagnetic fluorescence assay for pathogens.

In this work, we reported a simple rapid and point-of-care magnetic immunofluorescence assay for avian influenza virus (AIV) and developed a portable experimental setup equipped with an optical fiber spectrometer and a microfluidic device. We achieved the integration of immunomagnetic target capture, concentration, and fluorescence detection in the microfluidic chip. By optimizing flow rate and incubation time, we could get a limit of detection low up to 3.7 × 10(4) copy/µL with a sample consumption of 2 µL and a total assay time of less than 55 min. This approach had proved to possess high portability, fast analysis, high specificity, high precision, and reproducibility with an intra-assay variability of 2.87% and an interassay variability of 4.36%. As a whole, this microfluidic system may provide a powerful platform for the rapid detection of AIV and may be extended for detection of other viral pathogens; in addition, this portable experimental setup enables the development of point-of-care diagnostic systems while retaining adequate sensitivity.