RESUMO
OBJECTIVE: To describe an investigation of the feasibility of carrying out polymerase chain reaction (PCR) and fluorescent in situ hybridization on the same single human cell for sex determination. DESIGN: To develop protocol for a clinical diagnostic test in preclinical trials. SETTING: Infertile human volunteers in a clinical environment. PATIENTS: Polyploid embryos were obtained from patients treated by IVF at the Advanced Institute of Fertility. INTERVENTIONS: Seventeen blastomeres biopsied from human polyploid embryos were analyzed for sex determination by simultaneous application of PCR and fluorescent in situ hybridization. MAIN OUTCOME MEASURE: Presence of an X- or X and Y-chromosome band after agarose gel electrophoresis of PCR products and presence of an X- or X and Y-chromosome fluorescent probe signal after fluorescent in situ hybridization following PCR analysis. RESULTS: By PCR, all 17 blastomeres were amplified and, by fluorescent in situ hybridization, 12 (70.6%) blastomeres produced signals that were consistent with PCR results. Two blastomeres showed only X signals, although by PCR they had both X and Y-chromosome bands. CONCLUSIONS: The sequential use of PCR and fluorescent in situ hybridization on the same blastomere can be applied to improve the accuracy of sex determination before fresh ET.