Multiphoton microscopy also allows visualizing cells located underneath bone tissues such as cells of the bone marrow. Multiphoton microscopy provides considerably greater depth of penetration than single-photon confocal microscopy. If the area of interest is located more than 50–100 µm below the surface or there is a need to capture small-scale interactions between cells, multiphoton microscopy is required. The main consideration for the choice of a particular technique is the penetration depth needed to image the area and the amount of cell-cell interaction details required.
Intravital microscopy can be performed using several light microscopy techniques including widefield fluorescence, confocal, multiphoton, spinning disc microscopy and others. Microscope stage used for intravital microscopy imaging Cells need to produce a fluorescent protein to be visualized and this can be achieved by introducing a transgene. For example, in order to study the behavior of microglial cells in Alzheimer’s disease researchers will need to crossbreed a transgenic mouse that is a mouse model of Alzheimer’s disease with another transgenic mouse that is a mouse model for visualization of microglial cells.
The possibility of generating appropriate transgenic mice is crucial for intravital microscopy studies. Advancements in fluorescent protein technology and genetic tools that enable controlled expression of a given gene at a specific time in a tissue of interest also played important role in intravital microscopy development.
#VITAL IMAGING SOFTWARE#
High quality of modern microscopes and imaging software also permits subcellular imaging in live animals that in turn allows studying cell biology at molecular level in vivo. This is useful for many areas of research including immunology and stem cell research. Another advantage of intravital microscopy is that the experiment can be set up in a way to allow observing changes in a living tissue of an organism over a period of time. Thus, intravital microscopy allows researchers to study the behavior of cells in their natural environment or in vivo rather than in a cell culture. The main advantage of intravital microscopy is that it allows imaging living cells while they are in the true environment of a complex multicellular organism. Intravital microscopy involves imaging cells of a live animal through an imaging window that is implanted into the animal tissue during a special surgery. This technique is particularly useful to assess a progression of a disease or an effect of a drug. Intravital microscopy is used in several areas of research including neurology, immunology, stem cell studies and others. Animals are always anesthetized throughout surgeries and imaging sessions. Mice are the most common choice of animals for intravital microscopy but in special cases other rodents such as rats might be more suitable. For example, if researchers want to visualize liver cells of a live mouse they will implant an imaging window into mouse’s abdomen. īefore an animal can be used for intravital microscopy imaging it has to undergo a surgery involving implantation of an imaging window. Intravital microscopy is a form of microscopy that allows observing biological processes in live animals ( in vivo) at a high resolution that makes distinguishing between individual cells of a tissue possible. β-amyloid plaques (blue) are always present in brains of Alzheimer patients. The ability of microglial cells (green) to extend toward a laser lesion is reduced in Alzheimer’s disease mice. Microglial cells of this transgenic mouse produce GFP that allows cells to be visualised (green). Time-lapse two-photon intravital microscopy over a period of 54 minutes: microglial cells of the brain responding to acute laser lesion in an Alzheimer’s disease mouse.
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