Efficacy of calcium ionophore A23187 oocyte activation for generating parthenotes for human embryo research.
OBJECTIVE
Our purpose was to examine the efficacy of Ca-A23187 to activate human oocytes and produce parthenotes for research purposes. We examined the feasibility of using fluorescence in situ hybridization (FISH) to study the sex chromosome constitution of activated oocytes.


METHODS
One hundred eight nonfertilized oocytes from our IVF program were exposed to Ca-A23187. Oocyte activation was determined by the presence of pronuclear (PN) development. FISH was done on chromosome preparations using X and Y dual-colored probes. Polyploidic and parthenogenetically activated oocytes from our IVF program served as controls.


RESULTS
Of the 108 oocytes, 59 (55%) had no PN, 38 (35%) one PN, 10 (9%) two PN, and 1 (0.9%) three PN. Fiftyseven oocytes (53%) were not recovered following spreading and no chromatin was observed on 14 slides (13%) after FISH. This contrasted with 50 of 227 (22%) and 3 of 227 (1.7%) loss rates, respectively, for controls (P < 0.0001). Eight of 49 activated oocytes underwent cleavage. FISH was performed on 37 oocytes. Of 21 zero-PN oocytes, I had no FISH signals, 15 had a single X, 4 had two X's, and I had four X's. For one-PN oocytes, two had no FISH signals, seven had one X, and three had two X's. For two-PN oocytes, two had no FISH signals and two had two X's. FISH results were consistent with a maternal origin of genetic material.


CONCLUSIONS
Ca-A23187 resulted in a 45% activation rate, with 16% of oocytes progressing to cleavage before degeneration. Oocyte activation with Ca-A23187 allowed the generation of parthenotes for human embryo research. FISH was useful for evaluation of oocytes and parthenotes.
Rhoton-Vlasak
A
A
Department of Obstetrics and Gynecology, Mayo Clinic and Mayo Foundation, Rochester, Minnesota 55905, USA.
Lu
P Y
PY
Barud
K M
KM
Dewald
G W
GW
Hammitt
D G
DG
D001757:Blastomeres / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology*; D002455:Cell Division / Q000502:physiology; D002470:Cell Survival; D005456:Fluorescent Dyes; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism; D010312:Parthenogenesis / Q000502:physiology*; D012730:Sex Chromosomes
D005456:Fluorescent Dyes; D000001:Calcimycin; D002118:Calcium