prepare_samples
Differences
This shows you the differences between two versions of the page.
| Both sides previous revision Previous revision Next revision | Previous revision | ||
|
prepare_samples [2023/03/28 18:59] flowcytometry |
prepare_samples [2024/01/17 18:44] (current) flowcytometry [How do you get started?] |
||
|---|---|---|---|
| Line 4: | Line 4: | ||
| Talk to us about your experiment! If you want to use Flow Cytometry, we can help you design your new experiment. Get in contact with us and we will guide you through the Flow Cytometry Facility, and suggest the best systems for your project. Finally, we will book a training session with you and plan a trial run with your samples. | Talk to us about your experiment! If you want to use Flow Cytometry, we can help you design your new experiment. Get in contact with us and we will guide you through the Flow Cytometry Facility, and suggest the best systems for your project. Finally, we will book a training session with you and plan a trial run with your samples. | ||
| + | |||
| + | Here you can find summed up information on the essentials of handling your samples. | ||
| + | \\ | ||
| + | For more info check our **full document** on <html><a href="https://imm.medicina.ulisboa.pt/facility/flowcytometry/lib/exe/fetch.php?media=sop.ucf.005_-_how_to_prepare_samples_for_flow_cytometry.pdf" target="_blank"> How to prepare cell samples for Flow Cytometry</a></html>. | ||
| Line 21: | Line 25: | ||
| Notify us before bringing any cells of human or non-human primate origin to the core facility so we can help you assess the biosafety level of your cells. Most human primary cells and many human cell lines are BSL-2, posing hazards to laboratory staff and the environment.\\ Only BSL-1 and BSL-2 samples are allowed in the UCF room. BSL-3 and BSL-4 samples are not permitted. | Notify us before bringing any cells of human or non-human primate origin to the core facility so we can help you assess the biosafety level of your cells. Most human primary cells and many human cell lines are BSL-2, posing hazards to laboratory staff and the environment.\\ Only BSL-1 and BSL-2 samples are allowed in the UCF room. BSL-3 and BSL-4 samples are not permitted. | ||
| - | ***Analysis of cells by flow cytometry:** The analytical cytometers (BD LSRFortessa 1, 2, X-20, BD Accuri C6 and Amnis ImageStream Mark II) are suitable for BSL-1 work only. BSL-2 cells must be fixed before use on these instruments. If your work requires analysis of unfixed BSL-2 cells, this can be done on the BD FACSAria Fusion with aerosol containment. | + | ***Analysis of cells by flow cytometry:** The analytical cytometers (BD Accuri C6, BD LSRFortessa 2 / X-20, BD FACSymphony A5 SE, CYTEK Aurora and Amnis ImageStream Mark II) are suitable for BSL-1 work only. BSL-2 cells must be fixed before use on these instruments. If your work requires analysis of unfixed BSL-2 cells, this can be done on the BD FACSAria Fusion and BD FACSymphony S6 SE with aerosol containment. |
| - | ***Sorting of cells by flow cytometry:** BSL-2 unfixed cells (e.g. human samples) may only be sorted on the BD FACSAria Fusion with aerosol containment. BD FACSAria III can only sort BSL-1 and fixed BSL-2 material. | + | ***Sorting of cells by flow cytometry:** BSL-2 unfixed cells (e.g. human samples) may only be sorted on the BD FACSAria Fusion and BD FACSymphony S6 SE with aerosol containment. BD FACSAria III can only sort BSL-1 and fixed BSL-2 material. |
| Many staining protocols include a fixation and/or permeabilization step. However, if yours doesn’t and you need to fix your BSL-2 cells for analysis, we suggest **[[https://imm.medicina.ulisboa.pt/facility/flowcytometry/lib/exe/fetch.php?media=sop.ucf.005_-_how_to_prepare_samples_for_flow_cytometry.pdf|here]]** a quick and easy PFA fixation. | Many staining protocols include a fixation and/or permeabilization step. However, if yours doesn’t and you need to fix your BSL-2 cells for analysis, we suggest **[[https://imm.medicina.ulisboa.pt/facility/flowcytometry/lib/exe/fetch.php?media=sop.ucf.005_-_how_to_prepare_samples_for_flow_cytometry.pdf|here]]** a quick and easy PFA fixation. | ||
| Line 30: | Line 34: | ||
| **__Exclude your dead cells__** | **__Exclude your dead cells__** | ||
| - | Even though you keep your cells as happy as possible, some cells can die while handling and performing staining protocols. \\ Dead cells can affect the analysis by compromising the integrity of the data by non-specifically binding antibodies. That’s why most experiments benefit from adding a viability dye, to exclude dead cells from analysis and sorting. Also this viability staining can help you evaluate and compare the cell apoptosis and necrotic stages between cell conditions.\\ There are Fixable and Non-Fixable viability dyes. You can choose the right one for your assay depending on the purpose of your analysis and the post-staining protocol that will be performed. For more info check our full document on **[[https://imm.medicina.ulisboa.pt/facility/flowcytometry/lib/exe/fetch.php?media=sop.ucf.005_-_how_to_prepare_samples_for_flow_cytometry.pdf|How to prepare cell samples]]**. | + | Even though you keep your cells as happy as possible, some cells can die while handling and performing staining protocols. \\ Dead cells can affect the analysis by compromising the integrity of the data by non-specifically binding antibodies. That’s why most experiments benefit from adding a viability dye, to exclude dead cells from analysis and sorting. Also this viability staining can help you evaluate and compare the cell apoptosis and necrotic stages between cell conditions.\\ There are Fixable and Non-Fixable viability dyes. You can choose the right one for your assay depending on the purpose of your analysis and the post-staining protocol that will be performed. . |
| **__Perform the desired cell staining__** | **__Perform the desired cell staining__** | ||
prepare_samples.1680022757.txt.gz · Last modified: 2023/03/28 18:59 by flowcytometry
Page Tools
