sorting_faq
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sorting_faq [2018/03/05 18:02] flowcytometry |
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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 [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. | + | 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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