Table 1:

Comparison of super-resolution techniques and parameters that should be considered in choosing a super-resolution modality.

Technique	Commercial Availability	Equipment Cost	Lateral Spatial Resolution	Axial Spatial Resolution	Temporal Resolution	Sample penetration depth in standard setup	Specialized fluorophores	Difficulty of Data Analysis
STORM/PALM	High	Medium	~20–50 nm^a	~40–100 nm^b	Seconds-Minutes	Several μm	Yes	High
DNA-PAINT	Medium	Medium	~20–50 nm^a	~40–100 nm^b	Minutes-Hours^c	Several μm	No	High
ROSE/SIMFLUX	Low	Medium/High	~5–20 nm^a	N/A (implemented in 2D)	Minutes-Hours	Several μm	No^d	High
ROSE-Z	Low	Medium/High	~2–5 nm^a	~2–5 nm	Minutes-Hours	Several μm	No^d	High
ModLoc	Low	Medium/High	~3–5 nm^a	~7–10 nm	Minutes-Hours	Several μm	No^d	High
Expansion Microscopy	High	Low-Medium^e	~25–70 nm^f	~25–70 nm	Seconds-Minutes	Variable^g	No	Low
STED	High	High	~20–50 nm	~40–100 nm^h	10 ms-Minutesⁱ	Typically 10–15 μm, up to 120 μm^j	No	Low
RESOLFT	Medium	High	~20–50 nm	~40–100 nm	Seconds-Minutesⁱ	Several μm	Yes	Low
Parallelized RESOLFT	Low	High	~45–65 nm^k	~50–80 nm^k	Seconds-Minutes	Several μm	Yes	Low-Medium
MINFLUX	Medium	High	~1–3 nm	~1–3 nm	Down to 80 μsecondsⁱ	Several μm	Yes	High
SIM	High	Medium	~100–150 nm	~250–350 nm	Sub-seconds	Several μm^l	No	Medium
TIRF-SIM	High	Medium	~85–120 nm	~100–200 nm	Sub-seconds	200 nm	No	Medium
ISIM	High	Medium	~140–200 nm	~300–400 nm	Down to 10 ms	20 μm	No	Medium
Airyscan	High	Medium	~120–180 nm	~350–450 nm	Milliseconds-Secondsⁱ	Several μm	No	Low
SOFI	High	Low	~75–175 nm^m	~200–300 nm^m	Sub-seconds-Seconds	Several μm	Yes	High
SRRF	High	Low	~60–150 nm	Diffractionlimited (no improvement)	Seconds	Several μm	no	high

^a.

Values depend on many parameters, importantly the localization precision, which is dependent on the number of photons collected and the fluorophores used. Ranges correspond to most commonly used and bright fluorophores.

^b.

Values correspond to the most commonly used PSF engineering implementations (e.g. cylindrical lens) and can further be improved using interferometric approaches (iPALM, 4Pi-SMS and W-4Pi-SMSN)

^c.

Values are dependent on number of targets to image; with improved buffers and DNA-sequences this can be decreased to seconds-minutes

^d.

These methods can be implemented with STORM fluorophores or DNA-PAINT approaches. When implemented with DNA-PAINT, they do not require specialized fluorophores.

^e.

ExM can be done on a simple widefield microscope, but generally a confocal microscope is used due to sample size and thickness

^f.

70 nm refers to the most common ~4.5X sample expansion. With improved gel recipes or iterative expansions, larger expansions are possible, bringing resolution down to 25 nm

^g.

ExM imaging generally requires many Z-stacks based on increased sample size

^h.

Values correspond to most common implementation of 3D STED and can further be improved with the use of 4Pi geometry (isoSTED)

^i.

Temporal resolution inversely scales with the size of field of view.

^j.

Deep sample penetrance has been achieved with high N.A glycerol immersion lenses (Urban et al., 2011)

^k.

Lateral and axial resolution based on numbers reported in (Masullo et al., 2018) and (Bodén et al., 2021), respectively.

^l.

Up to 80 um depth has been demonstrated with adaptive optics (Lin et al., 2021).

^m.

The resolution here is based on 2^nd to 4^th order statistical analysis, the most common range for SOFI.