Disección del Midgut de una mosca adulta: un método para aislar el tracto digestivo
Published: April 30, 2023
Abstract
Fuente: Tauc, H.M., et al. Isolating Intestinal Stem Cells from Adult Drosophila Midguts by FACS to Study Stem Cell Behavior During Aging. J. Vis. (2014).
En este video, destacamos una disección del intestino adulto de Drosophila y la colección de midguts adecuados para la disociación unicelular y FACS con el fin de aislar las células madre intestinales.
Protocol
Este protocolo es un extracto de Tauc et al., Isolating Intestinal Stem Cells from Adult Drosophila Midguts por FACS to Study Stem Cell Behavior During Aging, J. Vis. Exp. (2014). NOTA: Si este protocolo se utiliza por primera vez para aislar los ISC por ordenación FAC, los siguientes controles son obligatorios para establecer inicialmente los parámetros FACS correctamente: Células disociadas del tipo salvaje (por ej…
Materials
| Forceps: Dumont, Inox Biologie #5 | Fine Science Tools | 11252-20 | |
| SefarNitex 03-150um/38 (35 µm nylon mesh) | Sefar | 3A03-0150-102-00 | |
| Falcon 5 ml Round Bottom Polystyrene Test Tube with Cell Strainer Snap Cap | Corning | 352235 | |
| Polymax 1040 | Heidolph | 543-42210-00 | |
| Albumin from bovine serum (BSA) | Sigma | A4503-50G | |
| 0.5% Trypsin-EDTA | Invitrogen | 15400-054 | Trypsin obtained from a different company most likely has a different activity and the duration of the trypsin digest has to be adjusted accordingly. |
| SYTOX Blue Dead Cell Stain for flow cytometry | Life Technologies | S34857 | |
| RNAlater Stabilization Solution | Life Technologies | AM7023 | Other solutions, e.g., Trizol can be used for subsequent RNA isolation |
| FACSAria II cell sorter | Becton Dickinson | Turn on one hour prior to sorting |
Cite This Article
Dissection of the Midgut from an Adult Fly: A Method for Isolating the Digestive Tract. J. Vis. Exp. (Pending Publication), e20150, doi: (2023).