Mesothelial Cell Isolation: A Technique to Isolate Primary Mesothelial Cells From Human Omentum Tissue Specimen
Published: April 30, 2023
Abstract
Source: Peters, P. N. et al. Modeling the Early Steps of Ovarian Cancer Dissemination in an Organotypic Culture of the Human Peritoneal Cavity. J. Vis. Exp. (2015)
This video describes the protocol to isolate primary human mesothelial cells from the human omentum specimen. The mesothelial cells can be used to establish an in vitro ovarian cancer model to investigate ovarian cancer cell adhesion, invasion, and proliferation.
Protocol
All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human welfare and have been reviewed by the local institutional review board. 1. Isolation and Culture of Primary Untransformed Stromal Cells Human tissue collection and preparation. Obtain specimens of human omentum, 2 cm3, removed during an abdominal surgery, and immediately immerse tissue in RT ph…
Representative Results
Figure 1. Isolation of primary human omental cells. (A) A piece of omentum is collected at surgery in PBS and transported to the laboratory. The tissue is pelleted at 0.5 x g x 3 min and transferred to fresh PBS. (B) The piece of omentum is transferred to a 10 cm petri dish, washed with PBS, and scalpels are used to cut t…
Açıklamalar
The authors have nothing to disclose.
Materials
| PBS | Fisher Scientific | SH3001304 | |
| Single-edged razor blades | Fisher Scientific | 12-640 | |
| 15 cm culture dishes | BD Biosciences | 353025 | |
| Fetal Bovine Serum (FBS) | Life Technologies | 16000044_3616914956 | |
| DMEM with L-Glutamine | Corning | 10-013-CV | |
| MEM Vitamins | Corning | 25-020-Cl | |
| MEM Nonessential amino acids | Corning | 25-025-CI | |
| Penicillin-Streptomycin | Corning | 30-002-CI | |
| Shaker | Thermo-Fisher | MaxQ 4450 | |
| Centrifuge | Eppendorf | 5702 | |
| Incubator | Thermo-Fisher | Forma Series II Water Jacketed CO2 Incubator Model 3100 | |
| T-75 Flasks | BD Biosciences | 353136 | |
| T-175 Flasks | BD Biosciences | 353112 | |
| Pipet tips | Rainin | P2, P10, P20, P200 and P1000 | |
| Microscope | Zeiss | Axiovert 200m | |
| Anti-Integrin αVβ3 Antibody, clone LM609 | EMD Millipore | MAB1976 | |
| Beta 1 | Oncosynergy | OS2966 | |
| Alpha 5 [CD49e] | ID Pharmingen | 555615 | |
| Beta 4 [CD104] | EMD Millipore | MAB 2058 | |
| Pipet tips | Corning | Filtered tips P2, P10, P20, P200 and P1000 | |
| Glass flask |
Etiketler
Bu Makaleden Alıntı Yapın
Mesothelial Cell Isolation: A Technique to Isolate Primary Mesothelial Cells From Human Omentum Tissue Specimen. J. Vis. Exp. (Pending Publication), e20376, doi: (2023).