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- Due to cell cycle asynchrony and tight adhesion between blastomeres, separation of early blastomeres must be done sequentially at each mitosis. For example, to isolate D blastomere, you will need to separate cell CD from cell AB at the first cell division, and then cell D from cell C at the division of the CD cell.
- Separation must be carried out on devitellinized embryos (with eyelash knife or hand-pulled fine glass wire). There is only a very brief window of time allowing you to separate two cells. All acts of separating cells must be carried out within a few minutes after the division has completed. Blastomeres quickly re-establish cell adhesion after division, and it would be impossible to separate two tightly adhered cells under such condition.
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