Table 1.

Comparison of the Amniotic Fluid Stem Cell-Extracellular Vesicles (AFSC-EVs) isolation techniques employed in the present study.

Technique	Manufacturer suggested amount	Method	Advantages	Disadvantages
UC	Regulated by centrifuge tube capacity	Pellet EVs at 100,000 g, after pre-clearing CM from cellular debris and live cells	High scalability (up to 32 mL when using Beckman Coulter rotor)	Inconsistent reproducibility across studies due to rotor size, UC time, speed and temperature Impurity of EV pellet due to aggregation of other particles Protocol may take >12 hours
ExoQuick	20% of CM	Reagent based methods that force precipitation of EVs out of solution due to water sequestration	Little processing time, but may require overnight incubation Expensive equipment not needed	Cost per preparation Retention of polymers in reagent
TEIR	50% of CM
Exo-PREP	100% of CM
qEV	500 µL of CM at a time, up to 4-time use	Sepharose beads in columns that fractionate CM based on gravity.	High yield of small size EV Low protein contamination Low sheer stress on EVs Fast protocol (15 minutes/preparation)	Column clogs and requires rinsing with NaOH/PBS to ensure adequate flow-through rate

UC: ultracentrifugation.

TEIR: Total Exosome Isolation Reagent.

CM: conditioned medium.

PBS: phosphate buffered saline.