Understanding the sub-cellular mechanisms in carcinogenesis is of crucial importance for cancer treatment. Popular cellular models comprise cancer cells grown as monolayers. But this approach disregards the three-dimensional (3D) interaction of tumor cells with their surrounding microenvironment. To understand the development and progression of malignancy in a close to nature context, the characterization of cancer microenvironments is crucial.
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A constantly ascending number of culturing methods allows cancer cells to be grown as 3D spheres. These cellular spheroids mimic the properties of solid tumors and are well suited to model cancer development with physiological relevance. The complex 3D structure and volumetric size represents a challenge in conventional light microscopy. This can be overcome by light sheet-based microscopy, which allows tracking of sub-cellular changes in large samples within reasonable time.
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The investigation of cellular spheroids with the Leica SP8 Digital LightSheet (DLS) microscope reveals meaningful results of cellular and molecular processes in standard petri dishes and is very well suited for cancer research.
Leica Microsystems’ Digital LightSheet module (DLS) can be integrated in the vertical axis of any inverted confocal Leica TCS SP8 microscope.
7.5 h time-lapse recording of mammary epithelial micro spheroid cultured in 3D. Data courtesy of intelligent imaging group (B. Eismann/C. Conrad) at BioQuant/DKFZ Heidelberg
In the past decade, the application of 3D cell cultures has grown significantly and has developed into a robust and reliable cellular model. More and more scientists are convinced that 3D cell cultures will soon replace conventional monolayer cell cultures.
Recent examples of 3D cell culture models elucidate new insights into colorectal cancer (CRC) and its complex underlying development mechanisms. In this case organoids show numerous valuable applications in disease research as well as in regenerative and personalized medicine.
The sample is prepared in standard glass bottom petri dishes. No special equipment, such as capillaries, is needed.
You can prepare several specimens within one experimental setup
The petri dish is turned upside down so that the spheroids sink into a position that guarantees optimal observability.
In the next step, the hydrogel solidifies at 37°C, producing a gel that will hold the spheroids in place.
Finally, the petri dish is turned upright again and filled with culturing medium.
Afterwards, the petri dish is put on the sample holder of the TCS SP8 DLS . The TwinFlect mirror provides 2-sided illumination for fast and easy imaging. Thanks to the low light exposure during image acquisition, it is possible to track the natural development of spheroids over long periods of time. The design of the DLS module enables you to view several samples within a single experiment (multi-position experiment). They can be scanned at more than 60 frames per second using the full chip of an sCMOS camera, one after the other, stack by stack, and over time. For convenient culture conditions you can add incubation systems of various manufactures, e.g. Okolab and TokaiHit.
Save your images as you acquire them by continuously streaming them from your temporary memory to your hard disk or a server solution (e.g. Acquifer HIVE).
Once the images are acquired, you can immediately visualize your results with the 3D Visualization tool of the Leica