Effect of enzymatic and mechanical methods of dissociation on neural progenitor cells derived from induced pluripotent stem cells

Abstract

Purpose To determine the most effective method of dissociating neural stem and progenitor cells into a single-cell suspension. Materials/methods Induced pluripotent stem cells were differentiated toward the neural fate for 4 weeks before clusters were subjected to enzymatic (Accutase, trypsin, TrypLE, dispase, or DNase I) or mechanical (trituration with pipettes of varying size) or combined dissociation. Images of cells were analyzed for cluster size using ImageJ. Results Cells treated with the enzymes Accutase, TrypLE, or trypsin/EDTA, these enzymes followed by trituration, or a combination one of these enzymes followed by incubation with another enzyme, including DNase I, were more likely to be dissociated into a single-cell suspension. Conclusions Cells treated with enzymes or combinations of methods were more likely to be dissociated into a single-cell suspension.

Systemic immunosuppression promotes survival and integration of subretinally implanted human ESC-derived photoreceptor precursors in dogs

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Novel adult cortical neuron processing and screening method illustrates sex- and age-dependent effects of pharmaceutical compounds

“…The dissociation of neurons from the extracellular matrix is an essential part of neuron isolation. Traditional methods include incubating the brain tissue in digestive enzymes 99 , digesting through mechanical means 100 , or both 101 . To maximize neuronal yield, we opted for using both methods.…”

Novel Adult Cortical Neuron Processing and Screening method illustrates Sex- and Age-dependent effects of pharmaceutical compounds

“…The dissociation of neurons from the extracellular matrix is an essential part of neuron isolation. Traditional methods include incubating the brain tissue in digestive enzymes 91 , digesting through mechanical means 92 , or both 93 . To maximize neuronal yield, we opted for using both methods.…”