Authors
Mohsen Norouzi, Seyedmajid Hosseini, Sydney Barker, Jose Cibelli, Jack C Koch, W Todd Monroe, Terrence R Tiersch, Jian Xu, Yue Liu
Published in
Lab on a chip. Nov 10, 2026. Epub Nov 10, 2026.
Abstract
Zebrafish embryonic cells are essential for a wide range of biotechnological and developmental biology applications including toxicology, disease modelling, genetic engineering, and genetic resource preservation. Efficient preparation of viable cells from zebrafish embryos requires removal of the protective chorion envelope followed by dissociation of embryonic tissues. Conventional approaches rely on manual manipulation or enzymatic treatment such as pronase which are labour-intensive, operator-dependent, costly, and require careful timing and handling to avoid embryo damage and experimental variability. We present for the first time a microfluidic platform that performs zebrafish embryo dechorionation and embryonic cell dissociation within an integrated device. The high-resolution 3D-printed dechorionation-cell dissociation (DCD) microfluidic device integrates mechanical dechorionation, graded micropost-based chorion capture, and flow-assisted embryonic cell dissociation in an enzyme-free, geometry-driven microfluidic system. Numerical simulations confirmed stable laminar flow (Re ≈ 13-344) throughout the device, with a moderate pressure drop (ΔP ≈ 3380 Pa at 1 mL s-1), suggesting that transient embryo blockage at geometric constrictions led to localized confinement-induced chorion rupture in the upstream dechorionation zone, while lower-stress downstream micropost regions (wall shear stress <1 Pa) supported subsequent dissociation of released embryonic fragments and cells. This separation of a geometrically confined dechorionation zone from a lower-stress downstream processing region enables effective chorion rupture while preserving cell viability. Experimental validation at 0.5-2.0 mL s-1 demonstrated reliable chorion rupture, trapping of chorions within the 500 μm micropost layers, and generation of embryo fragments and cells with sizes predominantly <20 μm (median), with 87% singlet events versus 89% for the manual method. Preprocessing and DCD processing took 8 s per embryo, compared with a reported 2 min per embryo for manual dechorionation alone. Cell viability ranged from 73-80%, approaching the manual method (84%), and processed cells remained viable and proliferative in culture for 14 d. A simple backflush rinsing protocol removed a substantial proportion of retained chorion material and supported short-term device reuse. The DCD microfluidic device provides a novel, scalable, reproducible, and enzyme-free microfluidic solution for automated zebrafish embryo processing with potential applications in somatic cell nuclear transfer, diploid germplasm cryopreservation, and biomedical research.
PMID:
43006645
Bibliographic data and abstract were imported from PubMed on 10 Nov 2026.
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