Stephanie L Angione1, Nathalie Oulhen2, Lynae M Brayboy3, Anubhav Tripathi1, Gary M Wessel4. 1. Center for Biomedical Engineering, School of Engineering, Brown University, Providence, Rhode Island. 2. Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island. 3. Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Women & Infants Hospital, Providence, Rhode Island; The Warren Alpert Medical School of Brown University, Providence, Rhode Island. 4. Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island. Electronic address: rhet@brown.edu.
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.
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.
Authors: Jin Akagi; Khashayar Khoshmanesh; Barbara Evans; Chris J Hall; Kathryn E Crosier; Jonathan M Cooper; Philip S Crosier; Donald Wlodkowic Journal: PLoS One Date: 2012-05-14 Impact factor: 3.240
Authors: Marcia A M M Ferraz; Heiko H W Henning; Tom A E Stout; Peter L A M Vos; Bart M Gadella Journal: Ann Biomed Eng Date: 2016-11-14 Impact factor: 3.934