Whole-Brain Optical Imaging

		Sample preparation/ slicing method	Typical voxel size	Data collection time	Suitable range of application	Whole-brain 3D dataset quality and consistency
Type	Typical method	Sample preparation/ slicing method	Typical voxel size	Data collection time	Suitable range of application	Whole-brain 3D dataset quality and consistency
LSFM	Ultramicroscopy 2007^［74］	Clearing	100-600 optical sections	6-8 h/mouse brain	Soma distribution，vascular network	★ Poor light sheet quality；inconsistent resolution，severe loss of deep quality
	Tiling LSM 2020^［80］	Clearing	2.6 μm×2.6 μm×6 μm（overall）， 0.3 μm×0.3 μm×1 μm（local）	5 h/mouse brain（2.6 μm× 2.6 μm×6 μm）	Soma distribution，vascular network，fine morphology of local neurons（axons，dendrites）	★★ Inconsistent resolution，loss of deep quality
	mLSFM 2021^［84］	Clearing，vibrating sectioning	0.8 μm×0.8 μm×5 μm（overall）， 0.3 μm×0.3 μm×1 μm（local）	95 min/mouse brain（0.8 μm×0.8 μm×5 μm）	Soma distribution，vascular network，fine morphology of local neurons（axons，dendrites）	★★ Inconsistent resolution；poor slicing performance and deformation
	VISoR 2021^［83］	Clearing	1 μm×1 μm× 2.5 μm	94 h（macaque brain slices）	Soma distribution，vascular network，neuronal axonal projections	★ Data loss between slices，difficult to register
STP	STP 2012^［86］	Agarose-embedding，vibrating sectioning	0.5 μm×0.5 μm×50 μm	24 h/mouse brain	Soma distribution，neuronal axonal projections	★★★ Consistent resolution and image quality；discontinuous Z-interval sampling
STP	MouseLight 2016^［87］	Clearing，vibrating sectioning	0.3 μm×0.3 μm×1 μm	8-10 days/mouse brain	Soma distribution，vascular and capillary network，complete fine morphology of neurons（axons，dendrites，spines）	★★★★ Poor slicing performance and deformation
FAST	FAST 2017^［91］	Agarose-embedding，vibrating sectioning	0.7 μm×0.7 μm×5 μm	2.4-10 h/mouse brain	Soma distribution，vascular network，neuronal axonal projections	★★ Consistent resolution and image quality
FAST	SMART 2021^［93］	Clearing，vibrating sectioning	0.7 μm×0.7 μm×30 μm（overall），0.3 μm×0.3 μm×1 μm（local）	20 h/part of mouse brain（0.3 μm× 0.3 μm×1 μm）	Soma distribution，vascular network，fine morphology of local neurons（axons，dendrites）	★★ Poor slicing performance and deformation；weak signals may go unrecognized and missing
MOST	MOST 2010^［94］	Nissl/Golgi staining，resin-embedded sectioning	0.33 μm× 0.33 μm×1 μm	242 h/mouse brain	Nissl stained soma，vascular and capillary network，Golgi stained neuron	★★ Consistent resolution and image quality；slight data loss between slice strips
	SI-fMOST 2016^［98］	Resin-embedded sectioning	0.32 μm× 0.32 μm×2 μm	3 days/mouse brain	Soma distribution，vascular and capillary network，complete fine morphology of neurons（axons，dendrites）	★★★★ Consistent resolution and image quality；natural self-registration，fine 3D reconstruction
	HD-fMOST 2021^［108］	Resin-embedded sectioning	0.32 μm× 0.32 μm×1 μm	111 h/mouse brain	Soma distribution，vascular and capillary network，complete fine morphology of neurons（axons，dendrites，spines）	★★★★★ Consistent resolution and image quality，high SBR；natural self-registration，fine 3D reconstruction
	Macaque brain imaging 2022^［109］	Hydrogel embedding，vibrating sectioning	0.65 μm× 0.65 μm×3 μm	70 days/macaque brain	Soma distribution，vascular network，neuronal axonal projections	★★★ Consistent resolution and image quality；natural self-registration，fine 3D reconstruction