An ultrasound-guided procedure to administer a label of DNA synthesis into fetal sheep

Abstract

A novel technique was developed to deliver a bolus dose of a DNA label into the peritoneal cavity of fetal sheep at 85–130 days gestation. Use of markers to identify the site of injection in fetuses from litters up to quadruplets, and immunohistochemistry to detect the DNA label, 5-bromo-2¢-deoxyuridine (BrdU), confirmed the procedure was successful in 85% of cases. Duration of the procedure was (mean SD) 44 16 min, and recovery from anaesthesia was rapid and uneventful in all cases. Fetal weight was estimated with a high degree of accuracy (residual standard deviation (RSD) = 297 g and r ² = 0.93, P<0.001) and the dose of label administered (110 33 mg BrdU/kg fetal weight) was adequate in all cases. BrdU detected in fetal nuclei following injection into amniotic fluid highlights the need for positive identification of the injection site in timed, short-term studies, and suggests potential to further develop the technique to investigate cellular events in fetal sheep younger than 85 days of gestation. The results demonstrate that the procedure can be used to determine in vivo whether or not nuclei have entered the S-phase of the cell cycle.