Hi everyone, I am going to spin an adherent MSC on object glass with cytospin. I will fix the cell with 4% PFA and do IF staining in the next month. What should I do to keep the fixed cell in an optimum condition?
Should I just keep it in slide box in -20 C or 4-10 C, should i soak with PBS or should I put inside a humid chamber?
Thank you.
Hi, To keep the fixed cells in an optimal state after 4% PFA fixation and before IF staining next month (I have stored it for up to 2 months and it still works with its problems of course), it is recommended to follow the following guidelines:
Sample storage: It is preferable to store fixed cell samples in a slide box under low temperature conditions. The optimal storage temperature to preserve the integrity of the fixed cells is generally at -20 °C for a maximum of 30 days or if it is going to take up to 3 months (I do not recommend more time) Store in a Deep Freezer at -70 C or colder. .
Protection against moisture: It is important to protect fixed cell samples from moisture to prevent the formation of ice crystals that can damage the cells. You can store the samples in a hermetically sealed slide box or suitable storage tubes to prevent exposure to moisture. If you choose to store in a slide box, make sure it is dry and properly sealed. Another technique that I sometimes use is after taking the samples, I give them a cold methanol bath, let them dry and finally store them, this with the objective that ice crystals do not form or as little as possible.
Storage medium: To avoid dehydration of the cells, you can soak fixed cell samples in a buffered buffered saline (PBS) solution before storing. This will help maintain proper humidity and prevent drying out. Make sure the samples are completely submerged in the PBS but under this method they don't last more than 15 days, hopefully 30 days.
Protection against light: It is advisable to protect fixed cell samples from direct exposure to light, especially bright light or ultraviolet light, as this can affect the integrity of the samples and compromise IF staining results, but if they are going to be in a freezer at -20 and covered with aluminum or a storage box with a cork bottom there should be no problem.
It is important to note that the optimal conditions may vary depending on the type of cells and the objectives of the specific IF staining. I have done the above with T lymphos, B lymphos and Mesenchymal cells, so I cannot assure you with other cell lines. .
Good luck in your experiments.
There is no problem in storing cell-containing coverslips in PBS (autoclaved) at 4°C for a month or even a bit longer. Just do not allow them to become contaminated by microorganisms.
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