The use of three-dimensional (3D) cell cultures for in vitro drug discovery assays has increased dramatically in recent years because 3D cell culture models more accurately mimic the in vivo environment compared to traditional two-dimensional (2D) monolayer cultures. However, current imaging-based analysis of these 3D cultures relies upon techniques originally developed for 2D cell culture, and as such, has significant limitations. Specifically, the light scattering inherent with thick microscopy specimens prevents imaging the entirety of a 3D spheroid, which are typically >100 µm in diameter. This technical limitation introduces a sampling bias in imaging analysis in which only the exterior cells of a spheroid can be imaged. To accurately survey the cellular environment and response of the spheroid, the field needs new techniques to overcome the sampling bias.
Visikol HISTO is an easy-to-use, rapid and non-destructive tissue clearing technique that is compatible with fluorescent protein, immunolabeling and various chemical dyes. Here we should its use with 1 mm brain tissue sections.
We are excited to announce that our 3Screen™ drug discovery services are available directly through both science exchange and scientist.com. Researchers at pharmaceutical companies can now easily leverage our digital pathology, 3D cell culture and whole mount imaging services for their drug discovery research questions.
Dr. Tom Villani from Visikol and Dr. Ann Rossi and Hilary Sherman from Corning published an article in Labcompare that discusses the challenges associated with imaging 3D cell culture models and extracting quantitative data.
With the widespread adoption of 3D cell cultures as more physiologically relevant in vitro models, there has arisen a need to adjust the methods of characterizing these models that may differ from standard 2D cell cultures. Neuronal spheroids are particularly challenging to image due to their dense nature and thus labeling methods need to be specifically optimized for their characterization.
Thermo Fisher Scientific recently launched the CX7 LZR HCS confocal system which has been integral to Visikol’s success in offering advanced 3D cell culture assays to its clients. The CX7 LZR system is ideal for 3D cell culture analysis as it is powered by seven solid state lasers which provide the ability to image 3D tissues past a depth of 1 mm when combined with Visikol® HISTO-M™ tissue clearing.
In order to maintain specimen integrity, one can utilize the high signal to noise benefits of fluorescent labeling along with tissue clearing and confocal microscopy to image full volumes of tissue to characterize blood vessels in three dimensions. Indeed, although the vasculature of various tissue types can form complex networks of interconnectivity, the labeling can be accomplished rather simply through a variety of techniques, ranging from simple use of highlighter ink[ii] to more specific immunolabeling such as with CD31[iii]. The technique implemented depends on the desired measurables, which may be as basic as total visualization of tissue vasculature, or quantitative comparison of functioning versus non-functioning blood vessels.
To help with slicing mouse brains into 1 mm thick sections, we have launched 1 mm coronal and sagittal mouse brain slicers which are inexpensive, easy-to-clean and easy to use. These slicers allow for rapid and easy tissue clearing and 3D tissue imaging.