Preparation, characterization and cell labelling of strong pH-controlled bicolor fluorescence carbonized polymer dots

Multicolor fluorescence N-doped CPDs from dextrin water solution in strong acidic and alkaline environments were synthesized and characterized, which revealed that pH value plays a vital role in the process of CPD growth.

Multicolor Fluorescence Imaging for the Early Detection of Salt Stress in Arabidopsis

Salt stress is one of the abiotic factors that causes adverse effects in plants and there is an urgent need to detect salt stress in plants as early as possible. Multicolor fluorescence imaging, as a powerful tool in plant phenotyping, can provide information about primary and secondary metabolism in plants to detect the responses of the plants exposed to stress in the early stage. The purpose of this study was to evaluate the potential of multicolor fluorescence imaging’s application in the early detection of salt stress in plants. In this study, the measurements were conducted on Arabidopsis and the multicolor fluorescence images were acquired at 440, 520, 690, and 740 nm with a self-developed imaging system consisting of a UV light-emitting diode (LED) panel for an excitation at 365 nm, a charge coupled device (CCD) camera, interference filters, and a computer. We developed a classification method using the imaging analysis of multicolor fluorescence based on principal component analysis (PCA) and a support vector machine (SVM). The results showed that the four principal fluorescence feature combinations were the ideal indicators as the inputs of the SVM model, and the classification accuracies of the control and salt-stress treatment at 5 days and 9 days were 92.65% and 98.53%, respectively. The results indicated that multicolor fluorescence imaging combined with PCA and SVM could act as a tool for early detection in salt-stressed plants.

A Novel Gesture-Based Control System for Fluorescence Volumetric Data in Virtual Reality

With the development of light microscopy, it is becoming increasingly easy to obtain detailed multicolor fluorescence volumetric data. The need for their appropriate visualization has become an integral part of fluorescence imaging. Virtual reality (VR) technology provides a new way of visualizing multidimensional image data or models so that the entire 3D structure can be intuitively observed, together with different object features or details on or within the object. With the need for imaging advanced volumetric data, demands for the control of virtual object properties are increasing; this happens especially for multicolor objects obtained by fluorescent microscopy. Existing solutions with universal VR controllers or software-based controllers with the need to define sufficient space for the user to manipulate data in VR are not usable in many practical applications. Therefore, we developed a custom gesture-based VR control system with a custom controller connected to the FluoRender visualization environment. A multitouch sensor disk was used for this purpose. Our control system may be a good choice for easier and more comfortable manipulation of virtual objects and their properties, especially using confocal microscopy, which is the most widely used technique for acquiring volumetric fluorescence data so far.

Snapshot multicolor fluorescence imaging using double multiplexing of excitation and emission on a single detector

Fluorescence-based multispectral imaging of rapidly moving or dynamic samples requires both fast two-dimensional data acquisition as well as sufficient spectral sensitivity for species separation. As the number of fluorophores in the experiment increases, meeting both these requirements becomes technically challenging. Although several solutions for fast imaging of multiple fluorophores exist, they all have one main restriction; they rely solely on spectrally resolving either the excitation- or the emission characteristics of the fluorophores. This inability directly limits how many fluorophores existing methods can simultaneously distinguish. Here we present a snapshot multispectral imaging approach that not only senses the excitation and emission characteristics of the probed fluorophores but also all cross term combinations of excitation and emission. To the best of the authors’ knowledge, this is the only snapshot multispectral imaging method that has this ability, allowing us to even sense and differentiate between light of equal wavelengths emitted from the same fluorescing species but where the signal components stem from different excitation sources. The current implementation of the technique allows us to simultaneously gather 24 different spectral images on a single detector, from which we demonstrate the ability to visualize and distinguish up to nine fluorophores within the visible wavelength range.

Coexpression of Three Odorant-Binding Protein Genes in the Foreleg Gustatory Sensilla of Swallowtail Butterfly Visualized by Multicolor FISH Analysis

Lepidopteran insects are mostly monophagous or oligophagous. Female butterflies distinguish their host plants by detecting a combination of specific phytochemicals through the gustatory sensilla densely distributed on their foreleg tarsi, thereby ensuring oviposition on appropriate host plants. In this study, to gain insight into the molecular mechanism underlying host plant recognition by the gustatory sensilla, using Asian swallowtail, Papilio xuthus, we focused on a family of small soluble ligand-binding molecules, odorant-binding proteins (OBPs), and found that three OBP genes showed enriched expression in the foreleg tarsus. Multicolor fluorescence in situ hybridization analyses demonstrated the coexpression of these three OBP genes at the bases of the foreleg gustatory sensilla. Further analyses on other appendages revealed that PxutOBP3 was exclusively expressed in the tissues which could have direct contact with the leaf surface, suggesting that this OBP gene specifically plays an important role in phytochemicals perception.

Cell culture-based enrichment of mouse hematopoietic stem and progenitor cells

Bone marrow (BM) chimeric mice are a valuable tool in the field of immunology and hematology, and genetic manipulation of donor cells is widely used to study gene function under physiological and pathological settings. Current BM chimera protocols generally require use of multicolor fluorescence-activated cell sorting (FACS) for donor hematopoietic stem and progenitor cell (HSPC) purification. Here, we describe a cell culture technique for the enrichment of functional HSPCs from mouse BM without the use of FACS purification. This technique is therefore expected to overcome current limitations in mouse BM chimera models.