Simple perfusion apparatus for manipulation, tracking, and study of oocytes and embryos.

ABSTRACT

OBJECTIVE: To develop and implement a device and protocol for oocyte analysis at a single cell level. The device must be capable of high resolution imaging, temperature control, perfusion of media, drugs, sperm, and immunolabeling reagents all at defined flow rates. Each oocyte and resultant embryo must remain spatially separated and defined. DESIGN: Experimental laboratory study. SETTING: University and academic center for reproductive medicine. PATIENT(S)/ANIMAL(S) Women with eggs retrieved for intracytoplasmic sperm injection (ICSI) cycles, adult female FVBN and B6C3F1 mouse strains, sea stars. INTERVENTION(S) Real-time, longitudinal imaging of oocytes after fluorescent labeling, insemination, and viability tests. MAIN OUTCOME MEASURE(S) Cell and embryo viability, immunolabeling efficiency, live cell endocytosis quantification, precise metrics of fertilization, and embryonic development. RESULT(S) Single oocytes were longitudinally imaged after significant changes in media, markers, endocytosis quantification, and development, all with supreme control by microfluidics. Cells remained viable, enclosed, and separate for precision measurements, repeatability, and imaging. CONCLUSION(S) We engineered a simple device to load, visualize, experiment, and effectively record individual oocytes and embryos without loss of cells. Prolonged incubation capabilities provide longitudinal studies without need for transfer and potential loss of cells. This simple perfusion apparatus provides for careful, precise, and flexible handling of precious samples facilitating clinical IVF approaches.