Analysis The Aneuploidy Of Frozen-thawed IVF Embryos From Older Women

Objective To detect the aneuploidy of the frozen-thawed IVF embryos from older women by FISH with fluorescent probes contained chromosomes 13,18,21, X and Y.Methods The frozen embryos after IVF-ET treatment were collected. Inclusion criteria:①Patients had been successful pregnancy and delivery after IVF-ET or FET treatment, and the couples had signed consent form with the intention to donate their remaining frozen embryos for scientific research;②Female infertility factors with the women age≥35 years;③There was no obvious chromosomal abnormalities with the couples. A total of 43 embryos from 7 couples were selected. All blastomeres of each embryo were acquired through biopsy after drilling a hole on the zona pellucida by OCTAX-LaserShot. The blastomeres were fixed using Tween-20/HCI +methanol/acetic acid. Fluorescence in situ hybridization processed with MultiVysion PGT probes (including chromosomes 13,18,21, X, Y five-color fluorescent probes). Then the FISH signals were analysised.Results A total of 43 embryos from 7 couples were selected.40 embryos were successfully revived after thawed. The recovery rate was 93.02%.2 embryos were excluded because in which there was only 1 alive cell. A totel of 270 blastomeres were alive in 41 embryos.264 intact blastomeres (97.78%) were acquired after biopsy.243 blastomeres(92.05%) in 264 blastomeres had nucleus,of which 12 blastomere had two nuclei and 2 blastomeres had 3 nuclei. 251 of 259(96.91%) nuclei were fixed successfully.247 of 251(98.41%) nuclei had FISH signals.9 embryos out of 41 were uniformly normal.32 embryos were abnormal, of which 2 non-mosaic embryos,5 chaotic embryos,25 mosaic embryos. There were 17 diploid mosaic embryos and 8 abnormal mosaic embryos embryos.11 embryos were uniform embryos,30 embryos were non-uniform embryos.11 embryos out of 41 had cells with multinuclei. All of those embryos were abnormal.22 out of 30 nuclei (73.33%) from 14 blastomeres were abnormal in cells with multinuclei. The abnormal rate was significantly higher than single-nuclei group (P<0.05). 103 out of 247 nuclei (41.70%) from 41 embryos nucleis were abnormal. The abnormal frequency of the chromosome 18 was the highest in the five chromosomes, followed by chromosome 13 and 21. The abnormal frequency of the sex chromosome was the lowest. In the diploid mosaic embryos, with the number of embryonic cells increased, the proportion of abnormal cells was decreased (rs=-0.520, P< 0.05).Conclusion1. High incidence of chromosomal abnormality existed in the frozen-thawed IVF embryos from older women. Mosiac was the main form.2. The abnormal frequency of the chromosome 18 was the highest in the frozen-thawed IVF embryos from older women.3. With the number of cells increased, the proportion of cells with chromosomal abnormality was decreased in the diploid mosaic embryos. There might be some self-repair mechanisms during human embryonic development.4. The rate of the chromosomal abnormality in multinuclei cells was significantly higher than single-nuclei cells.