sorting_faq
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sorting_faq [2018/03/05 16:08] flowcytometry [How are my cells collected?] |
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| - | ===== What should I expect when sorting? ===== | ||
| - | ==== Do I need to operate the equipment by myself? ==== | ||
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| - | We will train you to use the analyzers on your own. The sorters can only be operated by Flow Cytometry Facility staff. You can book time with us and we will sort for you. But if you want to learn cell sorting, we will teach you. | ||
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| - | ==== How are my cells collected? ==== | ||
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| - | {{ ::how_many_cells.png?400|}}You have many different options for collecting your sorted cells. | ||
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| - | * Collect up to four different populations simultaneously into tubes of your choice, 1.5 mL, 5 mL tubes, or up to two population into 15 mL tubes. | ||
| - | * Sort into any kind of plate. 6-well,12-well, 24-well,36-well, 96-well, 384-well, PCR plate, etc. | ||
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| - | Sort into the collection media of your choice. Collection tubes should be about 1/3 full of collection media. If you want to culture your sorted cells it’s a good idea to add Pen-Strep to the collection media. You can also sort directly into lysis buffer (for example, buffer that contains Trizol). | ||
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| - | We can adjust the temperature of the sorters to 4 ºC, 20 ºC, 37 ºC and 42 ºC, and we can also keep your cells on ice, if that is what you need. | ||
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| - | The BD FACSAria sorters faster with a smaller nozzle at higher pressure. With the 70 µm nozzle, you can sort up to 20,000 events/second. This is a great choice for small cells that are happy in single-cell suspension, for example blood and bone marrow. Larger cells need a larger nozzle size. Your cells should be no more than one-third the size of the nozzle. Cells that are fragile or easily stressed (e.g. cultured cells) need a lower pressure sort. Our sorters have the following options: | ||
| - | * 70 µm nozzle / 70 psi | ||
| - | * 85 µm nozzle / 45 psi | ||
| - | * 100 µm nozzle / 20 psi | ||
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| - | We normally use saline (0.9% NaCl) in our sorters as sheath fluid. Each individual cell is sorted inside a tiny drop of this sheath fluid. If you are using the 70 µm nozzle (high pressure, faster sorting), you will get approximately 1 ml of sheath fluid with every 1 million sorted cells. If you are using the 100um nozzle (low pressure, slower sorting), you will get approximately 3 ml of sheath fluid with every 1 million sorted cells. | ||
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| - | [[newuser|Back to New User]] | ||
sorting_faq.1520262496.txt.gz · Last modified: 2018/03/05 16:08 by flowcytometry
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