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Current safety data on chemotherapy during pregnancy are based on studies which focus on the mother and do not explore reproductive health and fecundity potential within the exposed offspring. We designed this randomized ex vivo animal study to evaluate the effect of chemotherapy on the developing ovarian reserve in the exposed offspring. Specimens (100 postnatal day zero C57BL/6 mouse ovaries) were randomized to control or chemotherapy drug exposure and maintained in a hanging well organ culture. Murine ovarian reserve establishment mirrors activity seen in the human fetus but with a significant time shift of the transition to meiotic arrest to the postnatal period. Exposures included: doxorubicin, cyclophosphamide, paclitaxel, docetaxel, and cisplatin. Doxorubicin resulted in a significant loss of 95.2% (p < 0.0001) of oocyte density compared to controls. Cyclophosphamide also caused depletion of 50.5% (p < 0.0001) of oocyte density. Cisplatin, docetaxel, and paclitaxel all demonstrated unique phenotypical changes on the ovaries and their oocytes, without a significant decrease in oocyte density over a five-day exposure. Exposure to chemotherapy may result in profound loss of oogonia during the transition to mature oocytes. [ABSTRACT FROM AUTHOR]