Labelled and label-free monitoring of immune cell proliferation
T-cell number and function can be examined via assays that detect T cell activity, including T-cell proliferation, cytokine production, and cytotoxicity. During infection, antigen-specific T lymphocytes undergo robust proliferation to form a large pool of effector T cells. This ability of T cells to proliferate in response to an antigen is measured by a lymphoproliferation assay and it has been used to indicate the presence of antigen-specific CD4+ helper T cells in a sample. To perform this assay, tested peripheral blood mononuclear cells (PBMCs) or purified T cells are mixed with the antigen, and the proliferation of cells is assessed using an endpoint assay. In contrast, live-cell imaging allows to monitor and quantify immune cell proliferation for hours or even days at a time, without missing any critical time points. Depending on the experimental set-up, both labelled and label-free approaches can be used to visualize and measure immune cell proliferation.
Visualization of dynamic immune responses
Phagocytosis is a key mechanism of the innate immune response that is used by neutrophils, macrophages, and other professional phagocytes to remove pathogens and cell debris from the body. To visualize and measure the process of phagocytosis, foreign material is tagged using pH-sensitive fluorescent probes that emit a signal upon exposure to the acidic environment of the phagosome. The CytoSMART Lux3 FL is a portable fluorescence live-cell imager that can track dynamic cellular processes, such as phagocytosis, in real time and under optimal cell culture conditions. In addition, the new Object Count algorithm of the CytoSMART Lux3 FL can automatically calculate the number of fluorescent objects in the image and investigate how this number changes over the course of time.
Real-time analysis of immune cell chemotaxis
The ability of certain immune cells, such as neutrophils and T cells, to detect an extracellular chemical signal and migrate in response to it, plays a crucial role in immune system function and homeostasis. The analysis of cell migration requires the application of high-throughput methods that can visualize and quantify cells’ motility in space and time. The CytoSMART live-cell imaging platforms, including the CytoSMART Omni and Lux3 FL, can be used for direct visualization and tracking of both, single-cell (e.g. macrophages, B and T lymphocytes) and collective cell migration (e.g. tumor cells). In case of single cell migration, the recorded time-lapse videos can be analyzed using commercially available (MetaMorph by BioImaging Solutions, Inc.) or free (TrackMate by ImageJ) software packages. Collective cell migration can be investigated using a wound-healing assay, with the changes in the rate and efficiency of cell movement quantified by means of the integrated CytoSMART image analysis software.