Estimation of the Mitochondrial Membrane Potential Using Fluorescence Lifetime Imaging Microscopy.

Monitoring of cell metabolism represents an important application area for fluorescence lifetime imaging microscopy (FLIM). In particular, assessment of mitochondrial membrane potential (MMP) in complex three-dimensional multicellular in vitro, ex vivo, and in vivo models would enable improved segmentation and functional discrimination of cell types, directly report on the mitochondrial function and complement the quenched-phosphorescence detection of cellular O and two-photon excited FLIM of endogenous NAD(P)H. Here, we report the green and orange-emitting fluorescent dyes SYTO and tetramethylrhodamine methyl ester (TMRM) as potential FLIM probes for MMP.

Application of Polyhydroxyalkanoates in Medicine and the Biological Activity of Natural Poly(3-Hydroxybutyrate).

Biodegradable and biocompatible polymers, polyhydroxyalkanoates (PHAs), are actively used in medicine to produce a wide range of medical devices and dosage formulations. The medical industry mainly utilizes PHAs obtained by chemical synthesis, but interest in the medical application of natural PHAs obtained biotechnologically is also growing. Synthetic PHAs are the biomimetic analogs of bacterial poly(3-hydroxybutyrate) (PHB) and other natural PHAs.

Circulating tumour cell RNA characterisation from colorectal cancer patient blood after inertial microfluidic enrichment.

The detection and molecular analysis of circulating tumour cells (CTCs) potentially provides a significant insight to the characterisation of disease, stage of progression and therapeutic options for cancer patients. Following on from the protocol by Warkiani et al. 2016, which describes a method of enriching CTCs from cancer patient blood with inertial microfluidics, we describe a method to measure the CTC RNA expression in the enriched fraction using droplet digital PCR and compare transcript detection with and without RNA pre-amplification.

A simple coating method of PDMS microchip with PTFE for synthesis of dexamethasone-encapsulated PLGA nanoparticles.

Dexamethasone is a widely used drug in medical and biological applications. Since the systematic and controllable release of this drug is of significant importance, encapsulation of this anti-inflammatory drug in poly(lactic-co-glycolic acid) (PLGA) nanoparticles can minimize uncontrolled issues. As dexamethasone-encapsulated PLGA nanoparticles are synthesized in the presence of organic solvents, poly(dimethylsiloxane) (PDMS)-based microchannels collapse due to the swelling problem.

Background-Free Fluorescence-Decay-Time Sensing and Imaging of pH with Highly Photostable Diazaoxotriangulenium Dyes.

Novel fluorescent diazaoxatriangulenium (DAOTA) pH indicators for lifetime-based self-referenced pH sensing are reported. The DAOTA dyes were decorated with phenolic-receptor groups inducing fluorescence quenching via a photoinduced-electron-transfer mechanism. Electron-withdrawing chlorine substituents ensure response in the most relevant pH range (apparent p K values of ∼5 and 7.

A Reappraisal of Circulating Fetal Cell Noninvasive Prenatal Testing.

New tools for higher-resolution fetal genome analysis including microarray and next-generation sequencing have revolutionized prenatal screening. This article provides commentary on this rapidly advancing field and a future perspective emphasizing circulating fetal cell (CFC) utility. Despite the tremendous technological challenges associated with their reliable and cost-effective isolation from maternal blood, CFCs have a strong potential to bridge the gap between the diagnostic sensitivity of invasive procedures and the desirable noninvasive nature of cell-free fetal DNA (cffDNA).

The Use of Microfluidic Technology for Cancer Applications and Liquid Biopsy.

There is growing awareness for the need of early diagnostic tools to aid in point-of-care testing in cancer. Tumor biopsy remains the conventional means in which to sample a tumor and often presents with challenges and associated risks. Therefore, alternative sources of tumor biomarkers is needed.

Cellulose-based scaffolds for fluorescence lifetime imaging-assisted tissue engineering.

Quantitative measurement of pH and metabolite gradients by microscopy is one of the challenges in the production of scaffold-grown organoids and multicellular aggregates. Herein, we used the cellulose-binding domain (CBD) of the Cellulomonas fimi CenA protein for designing biosensor scaffolds that allow measurement of pH and Ca gradients by fluorescence intensity and lifetime imaging (FLIM) detection modes. By fusing CBD with pH-sensitive enhanced cyan fluorescent protein (CBD-ECFP), we achieved efficient labeling of cellulose-based scaffolds based on nanofibrillar, bacterial cellulose, and decellularized plant materials.

A Novel Microfluidic Device-Based Neurite Outgrowth Inhibition Assay Reveals the Neurite Outgrowth-Promoting Activity of Tropomyosin Tpm3.1 in Hippocampal Neurons.

Overcoming neurite inhibition is integral for restoring neuronal connectivity after CNS injury. Actin dynamics are critical for neurite growth cone formation and extension. The tropomyosin family of proteins is a regarded as master regulator of actin dynamics.

Inertial-Based Filtration Method for Removal of Microcarriers from Mesenchymal Stem Cell Suspensions.

Rapidly evolving cell-based therapies towards clinical trials demand alternative approaches for efficient expansion of adherent cell types such as human mesenchymal stem cells (hMSCs). Using microcarriers (100-300 µm) in a stirred tank bioreactor offers considerably enhanced surface to volume ratio of culture environment. However, downstream purification of the harvested cell product needs to be addressed carefully due to distinctive features and fragility of these cell products.

Large-scale production of stem cells utilizing microcarriers: A biomaterials engineering perspective from academic research to commercialized products.

Human stem cells, including pluripotent, embryonic and mesenchymal, stem cells play pivotal roles in cell-based therapies. Over the past decades, various methods for expansion and differentiation of stem cells have been developed to satisfy the burgeoning clinical demands. One of the most widely endorsed technologies for producing large cell quantities is using microcarriers (MCs) in bioreactor culture systems.

Melanoma circulating tumor cells: Benefits and challenges required for clinical application.

The implementation of novel therapeutic interventions has improved the survival rates of melanoma patients with metastatic disease. Nonetheless, only 33% of treated cases exhibit long term responses. Circulating tumor cell (CTC) measurements are currently of clinical value in breast, prostate and colorectal cancers.

Development of Bright and Biocompatible Nanoruby and Its Application to Background-Free Time-Gated Imaging of G-Protein-Coupled Receptors.

At the forefront of developing fluorescent probes for biological imaging applications are enhancements aimed at increasing their brightness, contrast, and photostability, especially toward demanding applications of single-molecule detection. In comparison with existing probes, nanorubies exhibit unlimited photostability and a long emission lifetime (∼4 ms), which enable continuous imaging at single-particle sensitivity in highly scattering and fluorescent biological specimens. However, their wide application as fluorescence probes has so far been hindered by the absence of facile methods for scaled-up high-volume production and molecularly specific targeting.