Blastomere fixation techniques and risk of misdiagnosis for preimplantation genetic diagnosis of aneuploidy.
One of the most critical steps in preimplantation genetic diagnosis (PGD) studies is the fixation required to obtain good fluorescence in-situ hybridization (FISH) nuclear quality without losing any of the cells analysed. Different fixation techniques have been described. The aim of this study was to compare three fixation methods (1, acetic acid/methanol; 2, Tween 20; 3, Tween 20 and acetic acid/methanol) based on number of cells lost after fixation, average rate of informative cells, rate of signal overlaps and FISH errors. A total of 100, 106 and 114 blastomeres were fixed using techniques 1, 2 and 3 respectively. Technique 2 gave the poorest nuclear quality with higher cytoplasm, number of overlaps and FISH errors. Although technique 1 showed better nuclear quality in terms of greater nuclear diameter, fewer overlaps and FISH errors, it is difficult to perform correctly. However, technique 3 shows reasonably good nuclear quality and is both easier to learn and use for PGD studies than the others.
Velilla
Esther
E
Institute for Reproductive Medicine and Science of Saint Barnabas Medical Centre, 101 Old Short Hills Road, Suite 501, West Orange, NJ-07052, USA.
Escudero
Tomas
T
Munné
Santiago
S
D000782:Aneuploidy; D001757:Blastomeres / Q000502:physiology*; D003627:Data Interpretation, Statistical; D005307:Fertilization in Vitro; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D019836:Preimplantation Diagnosis; D016707:Tissue Fixation / Q000379:methods*