Tethering basics

Tethering station with a dissecting microscope, cooling platform, micromanipulator, and tethering tools Photo by Frank Loesche

To record a fly on the ball you first glue it to a small pin (the “tether”) so its head and body are held still while its legs walk freely. Tethering is a craft — the single biggest factor in getting clean data. A well-tethered, happy fly gives you hours of behavior; a crooked or over-glued one won’t walk.

The Fly Lab Gear tethering-station guide has the station design, hardware details, and source files.

The idea

  • Flies are cold-anesthetized on ice — no CO₂. Below about 4.5 °C, they stop moving within minutes; when they warm up, they recover and begin moving again. Keep them cold while you work, then let them warm to wake.
  • You glue only the top of the thorax to the tether. Head, legs, wings, and abdomen stay free.
  • Minimal glue. If you never lose a fly because it breaks free, you’re using too much glue.

What you need

Tethers (pins), a micromanipulator plus a Peltier-cooled tethering platform (the “sarcophagus”), light-curing glue plus a curing LED, a glass slide, a pin vice, a fine paintbrush, a vacuum pen, Kimwipes, and a dissecting scope.

Prepare the flies

  1. Get a vial; note the genotype and estimated age in your notebook.
  2. Transfer a few flies into a tube; plug it with cotton/foam. Every fly is precious — ask for small batches. A transfer funnel may help.
  3. Chill on ice until fully immobilized (~5 min). Do not leave them on ice longer than necessary: a longer chill means a longer recovery before the fly is ready to walk.

Prepare the station

  1. Power the station; confirm the platform is getting cold.
  2. Lay a small Kimwipe square on the cold sarcophagus to soak up condensation.
  3. Put a drop of light-curing glue on a glass slide.
  4. Mount a tether on the micromanipulator and practice moving it — smooth, controlled motion is half the battle.

Tether a fly

  1. Check the flies are immobile and the platform is very cold. Dab away any condensation with a twisted Kimwipe point.
  2. Scatter a few chilled flies onto the Kimwipe on the platform.
  3. Pick a good candidate: a large, healthy fly, completely still, wings folded back and legs tucked into a neat “hexagon.”
  4. With the vacuum pen, move it into a sarcophagus cavity, head pointing away from you. Nudge it with the paintbrush until the body is straight and you’re looking straight down on the thorax.
  5. With the pin vice, place a small drop of glue on the top ⅓ of the thorax (not on the head, wings, or leg joints).
  6. Lower the tether into the glue with the micromanipulator — touch the glue, don’t crush the fly.
  7. Cure the glue with the LED.
  8. Lift the fly off with the micromanipulator and set it on the waiting plate. It should wake and start moving within a minute.

Good vs. bad tether

✅ Good ❌ Bad
Tiny glue drop, thorax only Glue on legs, wings, joints, or head
Head free, body straight, level Fly cocked to one side / head glued
Wakes and walks within ~1 min Slow/no recovery (usually too much glue or too cold too long)

Mount on the ball

  1. Clamp the tether in the rig holder so the fly is centered over the ball.
  2. Lower until the legs just reach the ball and it grips and starts to walk.
  3. Confirm it walks: the ball turns under it, and once FicTrac is running you’ll see motion on the oscilloscope.

Practice first

Tethering takes reps — everyone should tether several practice flies before touching experimental ones. Time spent here pays off in every experiment after.

Reference

  • Loesche & Reiser (2021), An Inexpensive, High-Precision, Modular Spherical Treadmill Setup Optimized for Drosophila Experiments: doi:10.3389/fnbeh.2021.689573.

Updated 2026-07-10 01:47 ET.

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