Comparison of diSPIM With Confocal
The following comparison was written with diSPIM in mind, but the main points apply to all types of SPIM/LSFM.
Spinning disk confocal (SDCM) is just a massively parallel confocal implementation so it's faster but otherwise has the same characteristics as laser scanning confocal.
- Light dose: diSPIM has much lower dose and hence less photobleaching/phototoxicity than confocal. Reducing light dose is the main motivation for most SPIM users.
- A rule of thumb is that you need X times as much light for confocal as you do for SPIM where X is the number of slices in your stack.
- Figure 3 and Supplementary Video 2 of Wu et al. offer bleaching comparisons of imaging with diSPIM and SDCM. Other papers such as this one offer comparisons of light sheet with confocal which should apply to diSPIM.
- Z resolution: diSPIM after registration/fusion has best Z-resolution (same as XY resolution). Next best is confocal, which in turn has better Z resolution than single-view SPIM.
- Using Bessel beams to create the sheet can improve Z-resolution for single-view SPIM (“optical sectioning”), but won't meaningfully change dual view resolution after registration/fusion.
- XY resolution: confocal and SPIM are comparable.
- Speed: SPIM (per view) and SDCM have comparable speed if SDCM laser intensity is increased to compensate for the ~3% open area of the disks.
- Camera readout speed bounds the maximum achievable frame rate. For example, 512 pixels high ROI is 2.5 ms readout time for sCMOS. Allowing 2.5 ms illumination time results in 5 ms total per image or 200 frames per second. Using a cylindrical lens instead of a scanned light sheet can reduce camera overhead.