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Cell Biology JoVE (Open Access) Citable · DOI

In-vivo Centrifugation of Drosophila Embryos

DOI: 10.3791/2005-v
What you'll learn
  • ✓ Prepare and dechorionate Drosophila embryos for centrifugation
  • ✓ Mount live embryos in agar for density-based organelle separation
  • ✓ Separate organelles by centrifugal force along embryo axis
  • ✓ Recover and process embryo fractions for biochemical analysis
Protocol

We describe a method to separate organelles by density in living Drosophila embryos. Embryos are embedded in agar and centrifuged. This technique yields reproducible separation of major organelles along the anterior-posterior embryo axis. This method facilitates colocalization experiments and yields organelle fractions for biochemical analysis and transplantation experiments.

Difficulty
intermediate
Total time
~3–4 hours per cohort (including embryo collection, dechorionation, mounting, centrifugation, and recovery)
Model organism
Drosophila melanogaster (fruit fly embryos)
Biosafety
BSL-1

Steps

1
Prepare apple juice agar plates and harvest embryos

Set up apple juice agar plates as egg-laying substrate and collect Drosophila embryos at appropriate developmental stages for centrifugation experiments.

▶ 00:43
2
Remove chorion from embryos by dechorionation

Treat collected embryos with bleach solution to dissolve the protective egg shell (chorion), exposing the embryo for subsequent agar mounting.

▶ 03:26
3
Mount dechorionated embryos in agar

Embed prepared embryos into molten agar in a centrifuge tube, orienting them consistently along the anterior-posterior axis for uniform centrifugal separation.

▶ 05:09
4
Centrifuge embryos and recover organelle fractions

Spin agar-embedded embryos at defined conditions to stratify organelles by density, then dissect, homogenize, or section embryos to collect separated organelle fractions for downstream biochemical or transplantation analysis.

▶ 08:28
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