Experiment Overview
| Repository ID: | FR-FCM-Z2D4 | Experiment name: | HTFC screening protocol | MIFlowCyt score: | 97.58% |
| Primary researcher: | Annelisa Cornel | PI/manager: | Annelisa Cornel | Uploaded by: | Annelisa Cornel |
| Experiment dates: | 2019-07-25 - 2019-10-08 | Dataset uploaded: | Dec 2019 | Last updated: | Dec 2019 |
| Keywords: | [antibody staining] [high throughput flow cytometry] [drug screening] [adherent cells] [multiplex staining] | Manuscripts: | [31876358] |
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| Organizations: |
UMC Utrecht, Center translational immunology, Utrecht, Utrecht (The Netherlands)
UMC Utrecht, Center of Translational Immunology, Utrecht, Utrecht (Netherlands) |
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| Purpose: | To develop a ?no-touch? antibody staining method to be able to perform flow cytometry on adherent cells in a high-throughput fashion. | ||||
| Conclusion: | We show that we have generated a universal ‘no-touch’ antibody staining method to perform HTFC. The unmodified protocol works with several cell lines from different cellular origins. We show that multiplexing the staining does not influence the staining efficacy. The obtained results are in concordance with conventional FACS. | ||||
| Comments: | None | ||||
| Funding: | Not disclosed | ||||
| Quality control: | Fluorescent labelled beads (CS&T beads, Becton Dickinson) were used to check the performance and verify optical path and stream flow of the flow cytometer. This procedure enables controlled standardized results and allows the determination of long-term drifts and incidental changes within the flow cytometer. No changes were observed which could affect the results. In every experiment, appropriate controls have been taken into account: unstained, medium controls. Every condition, with every staining combination and every cell line has been performed at least in duplicate within individual experiments. Effects of TNF-α/IFN-γ are confirmed with ‘conventional’ flow cytometry. | ||||
Experiment variables
| Conditions | |
|---|---|
| · Medium treated | HLA-ABC stained medium control GIMEN 2.fcs · HLA-ABC stained medium control GIMEN.fcs · HLA-ABC stained medium control HEK.fcs · HLA-ABC stained medium control MCF-7.fcs · HLA-ABC stained medium control SKBR3 2.fcs · HLA-ABC stained medium control SKBR3.fcs · Unstained GIMEN 2.fcs · Unstained GIMEN.fcs · Unstained HeLa .fcs · Unstained MCF-7.fcs · quadriple stained medium control HEK 1.fcs · quadriple stained medium control HEK 2.fcs · quadriple stained medium control MCF-7.fcs · quadriple staining medium control HeLa 2.fcs · quadriple staining medium control HeLa.fcs · unstained SKBR3.fcs · quadriple staining medium control GIMEN 2.fcs · quadriple staining medium control GIMEN.fcs · HLA-ABC stained medium control HeLa 2.fcs · HLA-ABC stained medium control HeLa.fcs · HLA-ABC stained medium control HepG2 2.fcs · HLA-ABC stained medium control HepG2.fcs · Unstained HepG2.fcs |
| · TNF-α treated | HLA-ABC stained TNFa stimulated GIMEN 1 .fcs · HLA-ABC stained TNFa stimulated GIMEN 2 .fcs · HLA-ABC stained TNFa stimulated GIMEN 3.fcs · HLA-ABC stained TNFa stimulated HEK.fcs · HLA-ABC stained TNFa stimulated MCF-7 2.fcs · HLA-ABC stained TNFa stimulated MCF-7.fcs · HLA-ABC stained TNFa stimulated SKBR3 2.fcs · HLA-ABC stained TNFa stimulated SKBR3.fcs · quadriple stained TNFa stimulated HEK 1.fcs · quadriple stained TNFa stimulated HEK 2.fcs · quadriple stained TNFa stimulated MCF-7 2.fcs · quadriple stained TNFa stimulated MCF-7.fcs · quadriple staining TNFa stimulated GIMEN 2.fcs · quadriple staining TNFa stimulated GIMEN.fcs · HLA-ABC stained TNFa stimulated HeLa 2.fcs · HLA-ABC stained TNFa stimulated HeLa.fcs · HLA-ABC stained TNFa stimulated HepG2 2.fcs · HLA-ABC stained TNFa stimulated HepG2.fcs |
| · or TNF-α + IFN-γ treated cells | quadriple stained TNFa IFNy stimulated HEK 2.fcs · quadriple stained TNFa IFNy stimulated HEK.fcs · quadriple stained TNFa IFNy stimulated MCF-7 2.fcs · quadriple stained TNFa IFNy stimulated MCF-7.fcs · quadriple staining TNFa IFNy stimulated HeLa 2.fcs · quadriple staining TNFa IFNy stimulated HeLa.fcs · HLA-ABC stained TNFa IFNy stimulated HeLa .fcs · HLA-ABC stained TNFa IFNy stimulated HeLa.fcs · HLA-ABC stained TNFa IFNy stimulated HepG2 2.fcs · HLA-ABC stained TNFa IFNy stimulated HepG2.fcs |
