From seeing to believing: labelling strategies for in vivo cell-tracking experiments

Abstract

Intravital microscopy has become increasingly popular over the past few decades because it provides high-resolution and real-time information about complex biological processes. Technological advances that allow deeper penetration in live tissues, such as the development of confocal and two-photon microscopy, together with the generation of ever-new fluorophores that facilitate bright labelling of cells and tissue components have made imaging of vertebrate model organisms efficient and highly informative. Genetic manipulation leading to expression of fluorescent proteins is undoubtedly the labelling method of choice and has been used to visualize several cell types in vivo. This approach, however, can be technically challenging and time consuming. Over the years, several dyes have been developed to allow rapid, effective and bright ex vivo labelling of cells for subsequent transplantation and imaging. Here, we review and discuss the advantages and limitations of a number of strategies commonly used to label and track cells at high resolution in vivo in mouse and zebrafish, using fluorescence microscopy. While the quest for the perfect label is far from achieved, current reagents are valuable tools enabling the progress of biological discovery, so long as they are selected and used appropriately.

Up‐regulation of golgi α‐mannosidase IA and down‐regulation of golgi α‐mannosidase IC activates unfolded protein response during hepatocarcinogenesis

“…To observe the proliferation and migration ability of cells in vivo , cells were labeled with carboxyfluorescein succinimidyl ester (CFSE), an amine‐reactive fluorescent dye, and followed in embryos 30. The MAN1A1 ‐overexpressing 293T cell line and the MAN1C1 ‐overexpressing Hep3B cell line were labeled with CFSE and implanted in the yolks of 2‐day‐postfertilization zebrafish embryos.…”

Comparative suitability of CFDA‐SE and rhodamine 6G for in vivo assessment of leukocyte‐endothelium interactions

“…The same holds true for rhodamine 6G, the widely applied dye for in vivo labelling of leukocytes for IVM studies [7,8]. In contrast, carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) has been rarely applied for IVM investigations [9][10][11] even though it is discussed as an ideal dye for flow cytometry-based proliferation studies [5,6,12].…”

“…In the context of IVM investigations, several dyes for leukocyte labelling have been applied for the analysis of leukocyte recruitment. To this day, the ideal dye does not exist [6]. Acridine orange presents with limitations due to its accumulation in parenchymal tissue and its potential light-induced phototoxic effect that alters hemodynamics and cell adhesion.…”

Opening windows on new biology and disease mechanisms: development of real-time in vivo sensors

“…The recent development of fluorescent proteins with excitation and emission wavelengths approaching the infrared promises to expand the utility of these probes for in vivo use, owing to the enhanced tissue permeability of long-wavelength light [20]. In this Theme Issue, Progatzky et al [21] provide a detailed summary of commonly used fluorescent proteins, molecular dyes and reporters for in vivo applications, with a particular emphasis on the advantages and caveats of each system. Development of improved detection devices and techniques to facilitate in vivo use of these reporters will be necessary for widespread adoption of these sensors for clinical use, however.…”