Comparison of electrostatic and mechanical cell sorting with limited starting material

Area of Science:

• Biotechnology
• Cell Biology
• Immunology

Background:

• Fluorescence-activated cell sorting (FACS) is crucial for isolating cell populations in research.
• Standard FACS often uses high pressure and electrostatic deflection, potentially affecting cell homeostasis.
• Low-pressure, mechanical valve-based sorting offers an alternative approach.

Purpose of the Study:

• To compare the performance of high-pressure (BD FACSAria™ III) and low-pressure mechanical valve (Miltenyi MACSQuant® Tyto®) FACS technologies.
• To evaluate sorting metrics, cell viability, and post-sort cellular function.
• To assess the impact of different FACS technologies on blood cell populations.

Main Methods:

• Parallel sorting of peripheral and umbilical cord blood samples on BD FACSAria™ III and Miltenyi MACSQuant® Tyto® instruments.
• Manual magnetic-based cell sorting used as a reference.
• Assessment of sorting purity, viability, heat-shock protein HSPA1A expression, and T-cell proliferation potential post-sorting.

Main Results:

• Both FACS technologies effectively isolated blood cell types, including CD34+ cells, with minimal immediate impact on viability.
• The Miltenyi MACSQuant® Tyto® instrument exhibited lower variation in cell recovery and viability compared to the BD FACSAria™ III.
• Tyto-sorted T-cells showed significantly higher in vitro proliferation potential, suggesting less perturbation than Aria-sorted cells.

Conclusions:

• Both high-pressure and low-pressure FACS technologies can isolate blood cells from limited samples with good viability.
• The mechanical valve-based Tyto instrument appears to cause less cellular stress, as evidenced by superior T-cell proliferation.
• The Miltenyi MACSQuant® Tyto® offers a potentially gentler FACS method for sensitive cell populations.