Suzanne Cawood

The Centre of Reproductive and Genetic Health, London. UK

The finer points of trophectoderm biopsy

Preparing for blastocyst biopsy

Excellent blastocyst culture system

- KPIs (Blastocyst formation rate, utilisation rate, %d5 vs. d6 formation rate

Group/individual culture until d5. If group culture, separate blasts morning of d5/6 into individual culture

Zona drilling d3 – ensure hole size optimum (too large, risk of premature cell loss; too small, risk of ‘pinching’ of trophectoderm cells)

Biopsy in HEPES media. Make dishes 1 hour prior to procedure warmed at 37°C (non-gassed)

Ensure microscope stage heated to temperature which ensures droplet temperature is at 37 °C

Ensure optimum service is provided. Biopsy at d5AM/PM + d6 AM/PM is possible.

When and what to biopsy?

WHEN? Take into account how long the diagnosis will take. Vitryfying blastocysts post biopsy? Or aiming for fresh embryo

transfer?

WHAT? Hatching or Hatched blastocysts. Hatching more ideal

If herniating – trophectoderm cells will have to be ‘sucked’ from zona – more risk of lysis and damage to the embryo

ICM position

The procedure

Rapid technique

Minimise trauma/damage to both embryo and cells biopsied

Use blastocyst holding pipette

Common difficulties:

Positioning of embryo/pipettes (Hatched blastocysts, ICM hatching out)

Do not take too many cells! Remember image is 3D

Control at all times – both holding, suction and laser. Utilise foot pedal if possible.

Minimise number of laser shots – maximise shot value. Laser shots between the junction of trophectoderm cells

Post biopsy, cells to be washed (PBS-PBA) and tubed (hand-pipetted) into empty eppendorf tubes

Technique of excising cells Laser shot and tear cells

Fewer laser shots and ‘brushing’ the cells off using the holding pipette.

- Difficulty: Cells are sticky! - more likely to leave cells on holding/aspiration pipette (potential contamination so ensure pipettes changed if necessary)- this technique has to be used for completely hatched blastocysts