sorting_faq
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sorting_faq [2018/03/05 16:08] flowcytometry [How are my cells collected?] |
sorting_faq [2021/03/29 19:19] flowcytometry removed |
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| ==== Do I need to operate the equipment by myself? ==== | ==== Do I need to operate the equipment by myself? ==== | ||
| - | 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. | + | We will train you to use the analyzers on your own. The sorters can only be operated by [[about|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. |
| ==== How are my cells collected? ==== | ==== How are my cells collected? ==== | ||
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| * Sort into any kind of plate. 6-well,12-well, 24-well,36-well, 96-well, 384-well, PCR plate, etc. | * Sort into any kind of plate. 6-well,12-well, 24-well,36-well, 96-well, 384-well, PCR plate, etc. | ||
| - | 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). | + | The most recommended collection buffer is your **cell culture medium with 10% FBS** or some other serum. 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 antibiotics Pen-strep or gentamycin, and antifungal agents to the collection media. You can also sort directly into lysis buffer (for example, buffer that contains Trizol). |
| - | 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. | + | 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. |
| - | 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: | + | The BD FACSAria sorts 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 (100 µm nozzle). Our sorters have the following options: |
| * 70 µm nozzle / 70 psi | * 70 µm nozzle / 70 psi | ||
| * 85 µm nozzle / 45 psi | * 85 µm nozzle / 45 psi | ||
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