title: Highly reproducible quantification of apoptotic cells using micropatterned culture of neurons.
abstract: The quantification of apoptotic cells is an integral component of many cell-based assays in biological studies. However, current methods for quantifying apoptotic cells using conventional random cultures have shown great limitations, especially for the quantification of primary neurons. Randomly distributed neurons under primary culture conditions can lead to biased estimates, and vastly different estimates of cell numbers can be produced within the same experiment. In this study, we developed a simple, accurate, and reliable technique for quantifying apoptotic neurons by means of micropatterned cell cultures. A polydimethylsiloxane (PDMS) microstencil was used as a physical mask for micropatterning cell cultures, and primary granular neurons (GNs) were successfully cultured within the micropattern-confined regions and homogeneously distributed over the entire field of each pattern. As compared with the conventional method based on random cultures, the micropatterned culture method allowed for highly reproducible quantification of apoptotic cells. These results were also confirmed by using GNs derived from mice with neurodegeneration. We hope that this micropatterning method based on the use of a PDMS microstencil can overcome the technical obstacles existing in current biological studies and will serve as a powerful tool for facilitating the study of apoptosis-involved diseases.
authors.affiliation: Stem Cell Neuroplasticity Research Group, Kyungpook National University, Daegu 702-701, South Korea; Department of Laboratory Animal Medicine, Cell and Matrix Research Institute, College of Veterinary Medicine, Kyungpook National University, Daegu 702-701, South Korea.
authors.forename: Hyun
authors.initials: H
authors.lastname: Lee
authors.affiliation: School of Mechanical Engineering, Kyungpook National University, Daegu 702-701, South Korea.
authors.forename: Gyu Man
authors.initials: GM
authors.lastname: Kim
authors.affiliation: School of Mechanical Engineering, Kyungpook National University, Daegu 702-701, South Korea.
authors.forename: Jin Ho
authors.initials: JH
authors.lastname: Choi
authors.affiliation: Stem Cell Neuroplasticity Research Group, Kyungpook National University, Daegu 702-701, South Korea; Department of Physiology, Cell and Matrix Research Institute, School of Medicine, Kyungpook National University, Daegu 700-842, South Korea; Department of Biomedical Science, BK21 Plus KNU Biomedical Convergence Program, Kyungpook National University, Daegu 700-842, South Korea.
authors.forename: Jong Kil
authors.initials: JK
authors.lastname: Lee
authors.affiliation: Stem Cell Neuroplasticity Research Group, Kyungpook National University, Daegu 702-701, South Korea; Department of Physiology, Cell and Matrix Research Institute, School of Medicine, Kyungpook National University, Daegu 700-842, South Korea; Department of Biomedical Science, BK21 Plus KNU Biomedical Convergence Program, Kyungpook National University, Daegu 700-842, South Korea. Electronic address: jsbae@knu.ac.kr.
authors.forename: Jae-Sung
authors.initials: JS
authors.lastname: Bae
authors.affiliation: Stem Cell Neuroplasticity Research Group, Kyungpook National University, Daegu 702-701, South Korea; Department of Laboratory Animal Medicine, Cell and Matrix Research Institute, College of Veterinary Medicine, Kyungpook National University, Daegu 702-701, South Korea. Electronic address: hkjin@knu.ac.kr.
authors.forename: Hee Kyung
authors.initials: HK
authors.lastname: Jin
mesh_terms: D000818:Animals; D017209:Apoptosis; D001854:Bone Marrow Cells / Q000166:cytology; D002478:Cells, Cultured; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D059630:Mesenchymal Stem Cells / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D019636:Neurodegenerative Diseases / Q000378:metabolism / Q000473:pathology; D009474:Neurons / Q000166:cytology* / Q000378:metabolism
chemical_list: D004129:Dimethylpolysiloxanes; C013830:baysilon
keywords: Apoptosis; Granule neuron; Micropattern; Microstencil; Quantification