Table 2:

Comparison of the most frequently used sample delivery approaches

Delivery method	Crystal size	Mother liquor restrictions	Background scattering	Triggering compatibility	Technical Complexity	Efficient use of		Sample need for testing/static data set	Comments and Caveats
						sample	beamtime
Fixed targets/chips
SF-ROX (Goniometer mounted single crystals)	Several 100 μm	None. Typically cryocooled	low	Only electric field jumps	Low	High (but ~ 50 μm translations between exposures)	medium due to frequent crystal changes	Few crystals/50–100 crystals	SF-ROX, similar to helical scans at synchrotron sources
Fixed targets (Non-patterned chip)	Any size	None, LCP possible	Low for thin sample thickness and thin films	Pump probe (light scattering may be an issue) Slow chemical mixing in humidity-controlled environment	low	Lower than patterned	high	5 μl conc. cystal suspension/1–2 chips	Loading must be fast or in humid atmosphere to prevent sample dehydration which can result in nonisomorphism or other changes in diffraction properties. Samples and foils must be thin (few μm) for low background. Evaporation through thin foils or XFEL generated shot holes may be an issue, in particular when using vacuum chambers. The latter as well as diffusion of X-ray induced radicals affects spacing between exposures. Spacing between adjacent exposures in a row can be shorter than between rows. Frequent chip changes required (typically 10–15 min/chip)
Fixed targets (Patterned chip)	Should fit hole size	Not too viscous	Low for thin sample thickness and thin films	Pump probe (light scattering may be an issue in transparent chips); Chemical mixing in humidity-controlled environment long time delays possible	low	High	high	5–50 μl/1–2 chips	Loading and foil considerations as above. Moreover, blotting of mother liquor via perforated foils may result in crystal dehydration. The well depth determines the sample thickness (liquid film) in bottomless chips.
Droplet
DoD DoT (tape)	Big enough for good signal, 5–200 μm, but smaller than ID of capillary	Not too viscous (<40% PEG5000 or 35% glycerol)	High (due to droplet size and when hitting tape	Pump probe, chemical mixing 0.1 s - ~12s time delays	high	High	high	100 μl, ideally 30 % crystals (v/v) / 200 μl	ID of capillary sets upper limit on crystal size (Standard is 200 μm ID (which works fine for up to 80 μm longest crystal dimension)
Jets
GDVN	<20 μm	Not too viscous	Very low	Pump probe (fs-few μs) Chemical mixing, long time delays challenging	high	Very low	Very high, including MHz	300 μl, ideally 10–20 % crystals (v/v)/1–2 ml	Clogging of nozzles -> filtration of samples, prior and during injection, larger ID capillaries can help (50–100 μm) Settling of crystals -> anti-settling devices Fast video analysis to ensure jetting (and not spraying) Do not collect in the breakup region of the jet -> low, unreliable hit rate or very close to nozzle Test injection before beam time Compatible with MHz data collection
High viscosity extruders	Big enough to yield good signal but smaller than ID of the nozzle	None for grease-like matrix (but incompatible with some membrane protein crystals, causes dehydration in some crystals), LCP, cellulose, agarose, … limits on salt concentration, pH (not highly acidic)	high	Pump probe	Medium	High	high	5 μl/50 μl	ID of nozzle (50–100 μm) sets upper limit on crystal size Clogging Uneven flow rates, complicating in particular time-resolved experiments The flow can be disrupted by unattenuated XFEL beam Efficient sample use for XFELs with pulse repetition rate of 120 Hz and lower, as well as synchrotrons