Live-cell imaging is important for many different applications. However, its routine use has been constrained by the limitations of conventional live-cell imaging methods. This article comments on the growing popularity of automated live-cell imaging systems and highlights some key features to look for during platform selection.
Live-cell imaging is a microscopy-based technique used to examine living cells in real time. Compared to analyzing fixed samples, it offers deeper insights into dynamic cellular processes such as migration, development, and trafficking and can reveal findings that might otherwise be overlooked. Both bright field and fluorescence-based live-cell imaging modalities have been developed to support a range of different needs.
Applications of live-cell imaging span basic research through to biopharmaceutical manufacturing. In a research setting, live-cell imaging can be used during routine cell culture to help define the best time for subculture or harvest, determine the senescence status of cells, or detect early signs of contamination. Functional applications include studying cell migration (e.g., via a wound healing assay) and assessing experimental drug treatments for cytotoxicity or anti-proliferative effects. Where a fluorescence readout is employed, live-cell imaging can additionally be used for purposes that include evaluating transfection efficiency, following the production of reactive oxygen species, investigating apoptosis or phagocytosis, monitoring mitochondrial membrane potential, tracking calcium flux, and performing co-culture studies. Within the realm of biopharmaceutical manufacturing, live-cell imaging has broad utility for process development and control throughout the production of biologic drugs and vaccines.
Figure 1. Imaging Hela cells in apoptosis induced by staurosporine. Cells were treated with DEVD reagents conjugated to green-fluorescence to observe the activated caspase. Time-lapse images on both Bright field (upper images) and Green fluorescence channel (lower images) were collated using Celloger Nano. (Scale bar, 200 um)
Historically, live-cell imaging has involved manually monitoring cells by culturing them in a CO2 incubator and repeatedly removing the culture vessel to obtain images using a digital microscope. Drawbacks of this approach are that it is labor-intensive and highly prone to human error, largely because it offers no means of finding the same position in the culture vessel. Fluctuating environmental conditions can also cause cellular stresses, which can compromise results. While benchtop chamber devices improve on this method, they are bulky and cumbersome, and often struggle to maintain a stable environment.
Automated live-cell imaging systems address the inherent drawbacks of conventional technologies by being smaller, faster, more flexible, and easier to use. Depending on the intended application, factors to consider when selecting an automated live-cell imaging system include:
Compact live-cell imaging systems have been developed that fit inside a standard CO2 incubator. Not only do these platforms circumvent the need to purchase a dedicated incubator for live-cell imaging, but they are also designed to withstand the hot, humid conditions required to preserve cell viability.
Figure 2. Automated live-cell imaging systems, Celloger Mini Plus from Curiosis, placed inside a standard CO2 incubator
Live-cell imaging systems that offer both bright field and fluorescence options for either time-lapse or real-time monitoring offer maximum flexibility. To avoid cellular phototoxicity when measuring fluorescent readouts, it is important to select a system that demonstrates efficient performance at low light intensity.
Multipoint imaging (the measurement of multiple points per well or per vessel) can help to improve the statistical significance of results. Where multipoint imaging can be combined with data collection according to a set schedule (intervals, cycles, total time), it is especially useful for such applications as studying the effects of different drug concentrations on cells over time.
Live-cell imaging systems that are supplied with interchangeable holders for different vessel types (well-plates, culture flasks, culture dishes) are readily adapted to any workflow, especially where vessel specifications can be customized by brand, and can provide opportunities for scale up where required.
While some live-cell imaging systems employ a moveable stage, those featuring internal optics avoid perturbing the sample to ensure a more consistent environment. Stable scanning performance is further assured where the illumination can be precisely controlled by adjusting the gain, intensity, and offset value.
Ease-of-use is often a major deciding factor when purchasing laboratory equipment. Features to look for in an automated live-cell imaging system include whether it is equipped with autofocusing and if a Z-stacking function is available, such as is required for observing spheroid cells. The types of analysis possible should also be investigated; these might include intensity histograms, confluency graphs, and image stitching for high-resolution mapping of a large sample area.
Figure 3. Cell confluency analysis using Celloger Mini Plus analysis software. Images show the progress of confluency analysis of time-lapse images & cell confluence graph
The Celloger Mini Plus is an automated live-cell imaging system that fits inside a standard CO2 incubator for long-term cellular monitoring or time-lapse imaging. To learn more, visit curiosis.com