Structured illumination microscopy
02 Oct 2013
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Details regarding Structured illumination microscopy

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​Zeiss Elyra SIM/STORM microscope

 

Contacts: Dr. Lin Wang

Structured illumination microscopy

Structured illumination microscopy (SIM) is a method of imaging fluorescently labelled structures at greater spatial resolution than a conventional fluorescence microscope is capable of Optical diffraction imposes a limit of ~200 nm lateral and 500 nm axial resolution in the conventional microscope, both of which are doubled in SIM. It involves imaging a grating onto the sample to produce Moiré fringes, which may be resolved in the microscope even if the sample structure cannot be seen directly.

Why use Structured illumination microscopy?
As with other super-resolution techniques, such as single molecule localisation microscopy (SMLM) and stimulated emission depletion microscopy (STED), structured illumination microscopy (SIM) enables fine structure to be resolved. The advantage of SIM over other super-resolution imaging techniques is that the choice of fluorescent dye is limited solely by the laser lines available to excite it. It also uses much lower laser powers and hence is suited to applications where limiting phototoxicity is important or for time-lapse imaging. It is compatible with multicolour imaging of up to four fluorescent probes.
SIM can be performed on the same microscope as SMLM, enabling correlative imaging with the two super-resolution methods.

Applications
SIM is well-suited to imaging live cells because it has low phototoxicity and the imaging duration is low, of the order of seconds. It is equally applicable to animal and plant biology, as well as in chemistry and materials science.

Microscope Specification

 Instrument

  Zeiss Elyra PS1

 Laser wavelengths (nm)

  405, 488, 532, 561, 642

 Excitation power density

 

 Compatible objective lenses

  x100 oil immersion, x63 oil immersion, x63 water immersion

 Compatible dyes

  Any, subject to available laser wavelengths

 Compatible sample mounts

  #1.5 cover slip on microscope slide or glass bottom Petri dish

 Configuration

  Inverted microscope

 Temperature control

  21-37°C

 Lateral resolution

  ~100 nm (depends on wavelength)

 Axial resolution

  ~250 nm (depends on wavelength)

 Temporal resolution

  ~0.1 Hz for single plane

 Grating angles

  3 or 5

Case Studies

Characterisation of fluorescent nanomaterials - University of Bristol (link opens in a new window)

 

Contact: Wang, Lin (STFC,RAL,CLF)