FISH analysis of six chromosomes in unfertilized human oocytes after polar body removal.
OBJECTIVE
To develop an improved technique for estimating chromosomal abnormalities in human oocytes by fluorescence in situ hybridization (FISH) and to correlate the position of single chromatids with the chromosomal status of the oocytes.


METHODS
Oocytes that were at metaphase II about 17-20 hr after insemination or intracytoplasmic sperm injection (ICSI) were treated with pronase to remove the zona pellucida and polar body (PB) and then spread on slides using HCl and Tween 20. Two rounds of FISH were performed using direct-labeled probes: chromosomes 1, 13, 21 (round 1); chromosomes X, 7, 18 (round 2).


RESULTS
Of the 63 oocytes from 18 patients (mean age, 32 years), 48 (76%) had one DNA complement as expected, 9 (14%) had 2 DNA complements, 3 (5%) gave incomplete FISH signals, and 3 (5%) were not analyzable. Of the 48 oocytes with one set of DNA, 48% were haploid, 44% were aneuploid for one or more chromosomes, and 8% were polyploid. We also found an increased frequency of predivision of chromatid bivalents in aneuploid oocytes, especially for chromosome 21.


CONCLUSIONS
This technique enables simultaneous assessment of six chromosomes in human oocytes, and therefore can be useful for accurately determining the incidence and causes of genetic imbalances in human oocytes and apparently low fertilization rates.
Martini
E
E
Department of Molecular Cell Biology and Genetics, University of Maastricht, The Netherlands.
Flaherty
S P
SP
Swann
N J
NJ
Matthews
C D
CD
Ramaekers
F C
FC
Geraedts
J P
JP
D000328:Adult; D000782:Aneuploidy; D022003:Cell Nucleus Structures; D002842:Chromatids / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000032:analysis; D015342:DNA Probes; D005260:Female; D005306:Fertilization; D006238:Haploidy; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007211:Indoles; D009865:Oocytes / Q000166:cytology* / Q000378:metabolism / Q000473:pathology*; D011123:Polyploidy; D011402:Pronase / Q000378:metabolism; D020554:Sperm Injections, Intracytoplasmic
D015342:DNA Probes; D007211:Indoles; C007293:DAPI; D004247:DNA; D011402:Pronase