Light sheet-based fluorescence microscopy (LSFM) for the quantitative imaging of cells and tissues
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In light sheet-based fluorescence microscopy (LSFM), only the focal plane is illuminated by a laser light sheet. Hence, only the fluorophores within a thin volume of the specimen are excited. This reduces photo-bleaching and photo-toxic effects by several orders of magnitude compared with any other form of microscopy. Therefore, LSFM (aka single/selective-plane illumination microscopy [SPIM] or digitally scanned light sheet microscopy [DSLM]) is the technique of choice for the three-dimensional imaging of live or fixed and of small or large three-dimensional specimens. The parallel recording of millions of pixels with modern cameras provides an extremely fast acquisition speed. Recent developments address the penetration depth, the resolution and the recording speed of LSFM. The impact of LSFM on research areas such as three-dimensional cell cultures, neurosciences, plant biology and developmental biology is increasing at a rapid pace. The development of high-throughput LSFM is the next leap forward, allowing the application of LSFM in toxicology and drug discovery screening.
KeywordsLight sheet-based fluorescence microscopy (LSFM) Digitally scanned light sheet-based microscopy Single/selective-plane illumination microscopy Three-dimensional cell cultures Cellular spheroids High-throughput LSFM
We thank Nariman Ansari for many discussions on 3D cell culture, Daniel von Wangenheim for the comments on LSFM applications in plant research and Christian Mattheyer for his contribution to optical clearing.
- Dobosz M, Ntziachristos V, Scheuer W, Strobel S (2014) Multispectral fluorescence ultramicroscopy: three-dimensional visualization and automatic quantification of tumor morphology, drug penetration, and antiangiogenic treatment response. Neoplasia 16:1–W7. doi: 10.1593/neo.131848 PubMedCentralPubMedGoogle Scholar
- McLachlan D Jr (1968) Microscope. US. Retrieved from https://www.google.com/patents/US3398634?dq=D.+McLachlan+j.+microscope&hl=en&sa=X&ei=na8HVMh9wbI8_uWBoAI&ved=0CCIQ6AEwAA
- Olarte OE, Licea-Rodriguez J, Palero JA, Gualda EJ, Artigas D, Mayer J, Loza-Alvarez P (2012) Image formation by linear and nonlinear digital scanned light-sheet fluorescence microscopy with Gaussian and Bessel beam profiles. Biomed Opt Express 3:1492–1505. doi: 10.1364/BOE.3.001492 PubMedCentralPubMedGoogle Scholar
- Pampaloni F, Stelzer EHK, Mattheyer C (2014b) Kapillarzelle, anordnung und verfahren zur aufnahme, zur positionierung und zur untersuchung einer mikroskopischen probe. Retrieved from https://www.google.com/patents/WO2014033320A1?cl=de&dq=francesco+pampaloni&hl=en&sa=X&ei=_yIUVMjOHcHMyAPRq4GwDQ&ved=0CDsQ6AEwBA
- Wu Y, Ghitani A, Christensen R, Santella A, Du Z, Rondeau G, Shroff H (2011) Inverted selective plane illumination microscopy (iSPIM) enables coupled cell identity lineaging and neurodevelopmental imaging in Caenorhabditis elegans. Proc Natl Acad Sci U S A 108:17708–17713. doi: 10.1073/pnas.1108494108 PubMedCentralPubMedGoogle Scholar