Isolation of chicken primordial germ cells using fluorescence-activated cell sorting

Area of Science:

• Developmental Biology
• Cell Biology
• Genetics

Background:

• Germ line modification in chickens is hindered by difficulties in isolating primordial germ cells (PGC).
• Efficient fractionation of PGC from somatic cells is crucial for genetic manipulation and research.

Purpose of the Study:

• To develop a method for isolating an enriched population of viable chicken PGC.
• To enable efficient PGC isolation from embryonic blood and gonadal tissues for future germ line modification studies.

Main Methods:

• Harvested blood from early chick embryos (stages 13-15) and cells from gonads of stage 27 embryos.
• Labeled viable PGC with anti-stage-specific embryonic antigen-1 (SSEA-1) and sorted using fluorescence-activated cell sorting (FACS).
• Confirmed PGC identity using periodic acid-Schiff (PAS) or anti-embryonic mouse antigen-1 (EMA-1) staining.

Main Results:

• Isolated SSEA-1-positive cells represented <0.1% of blood cells and ~2% of gonadal cells.
• Recovery rates of sorted PGC on slides ranged from 53-73%, with high purity (92-94% positive for PAS or EMA-1).
• Sorted PGC demonstrated viability in vitro after isolation.

Conclusions:

• A reliable method for labeling and sorting viable chicken PGC from embryonic tissues using SSEA-1 and FACS has been established.
• This technique facilitates the isolation of PGC, paving the way for advancements in chicken germ line modification and genetic research.