Agarose Spin Down Assay: A Technique to Embed Organoids for Histological Analysis
Published: April 30, 2023
Abstract
Source: Lee, S. et al. Establishment and Analysis of Three-Dimensional (3D) Organoids Derived from Patient Prostate Cancer Bone Metastasis Specimens and their Xenografts. J. Vis. Exp. (2020)
In this video, we perform a spin-down assay to embed 3D prostate cancer organoids in agarose. These embedded organoids can be used for further histopathological analysis to study cancer growth and development.
Protocol
1. Organoid Processing for Histology: The Agarose Spin Down Method NOTE: This protocol is adapted from a previous publication by Vlachogiannis et al. We have added a step involving agarose embedding to embed all populations of organoids successfully. Remove existing media from the well. Be careful not to aspirate the basement membrane domes. Add an equal (equal to the volume of media removed from step 1) volume of cell recovery s…
Representative Results
Figure 1: A key step for successful agarose embedding. A process to detach the cell pellet from the wall of the 1.5 mL tube.
Divulgaciones
The authors have nothing to disclose.
Materials
| 1 mL Pipettman | Gilson | F123602 | |
| 1 mL Syringe | BD Syringe | 329654 | |
| 1.5 mL tube | Spectrum Lab Products | 941-11326-ATP083 | |
| 25G Needle | BD PrecisionGlide Needle | 305122 | |
| 4% Paraformaldehyde (PFA) | Alfa Aesar | J61899 | |
| 70% Ethanol (EtOH) | VWR | BDH1164-4LP | |
| Agarose | Lonza | 50000 | |
| Cell Culture Plate – 24 well | Costar | 3524 | |
| Cell Culture Plate – 48 well | Costar | 3548 | |
| Cell Culture Plate – 6 well | Costar | 3516 | |
| Cell Dissociation Solution, Accumax | Innovative Cell Technologies, Inc. | AM105 | |
| Cell Recovery Solution | Corning | 354253 | |
| Cell Scraper | Sarstedt | 83.18 | |
| dPBS | Corning/Cellgro | 21-031-CV | |
| Pipette tips for 1 mL (Blue Tips) | Fisherbrand Redi-Tip | 21-197-85 | |
| Small Table Top Centrifuge | ThermoFisher Scientific | 75002426 | |
| OCT Compound | Tissue-Tek | 4583 | |
| Water Bath | Fisher Sc | 2320 |
Citar este artículo
Agarose Spin Down Assay: A Technique to Embed Organoids for Histological Analysis. J. Vis. Exp. (Pending Publication), e20388, doi: (2023).