Abstract
We present spatial light interference microscopy (SLIM) as a new optical microscopy technique, capable of measuring nanoscale structures and dynamics in live cells via interferometry. SLIM combines two classic ideas in light imaging: Zernike’s phase contrast microscopy, which renders high contrast intensity images of transparent specimens, and Gabor’s holography, where the phase information from the object is recorded. Thus, SLIM reveals the intrinsic contrast of cell structures and, in addition, renders quantitative optical path-length maps across the sample. The resulting topographic accuracy is comparable to that of atomic force microscopy, while the acquisition speed is 1,000 times higher. We illustrate the novel insight into cell dynamics via SLIM by experiments on primary cell cultures from the rat brain. SLIM is implemented as an add-on module to an existing phase contrast microscope, which may prove instrumental in impacting the light microscopy field at a large scale.
Highlights
Most living cells do not absorb or scatter light significantly, i.e. they are essentially transparent, or phase objects
Phase contrast microscopy proposed by Zernike represented a major advance in intrinsic contrast imaging, as it revealed inner details of transparent structures without staining or tagging [1]
In order to assess the spatial accuracy of spatial light interference microscopy (SLIM), we imaged an amorphous carbon film deposited on glass and compared the topography measurements against atomic force microscopy (AFM)
Summary
Most living cells do not absorb or scatter light significantly, i.e. they are essentially transparent, or phase objects. QPI-based projection tomography has been applied to live cells [17,18,19] Despite these significant technological advances, the range of QPI applications in biology has been largely limited to red blood cell imaging [20,21,22] or assessment of global cell parameters such as dry mass [4,23], average refractive index [24], and statistical parameters of tissue slices [25]. This limitation is due to two main reasons, as follows. The SLIM image is intrinsically registered with the other channels of the microscope, including fluorescence, which enables powerful multimodal investigations
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