This article describes a method for isolating oligodendrocyte-conditioned medium (OCM) from oligodendrocyte lineage cells. The process enhances cell attachment and promotes the secretion of bioactive molecules that are crucial for various research applications.
Take a polymer-coated petri dish.
Add oligodendrocyte lineage, or OL cells to a suitable culture medium and incubate.
The positive charge on the polymer coating attracts the negatively charged cell membrane, enhancing cell attachment to the dish. The growth factors and nutrients in the medium facilitate cell expansion.
Replace the medium with a nutrient-poor medium and incubate.
Low nutrient levels induce mild stress in the cells, causing them to secrete proteins, lipids, and other molecules, including growth factors and cytokines, into the extracellular space.
Collect the medium in a tube.
Filter sterilize the medium to remove cellular debris and insoluble components.
The filtrate containing the soluble bioactive molecules secreted by OL cells is called oligodendrocyte-conditioned medium or OCM.
Store the OCM at lower temperatures until further use to prevent the degradation of proteins and other molecules, thereby preserving the effectiveness of the OCM.
Plate two or three pre-coated 100-millimeter Petri dishes with 10 milliliters of the cell suspension.
Incubate in a humidified incubator at 37 degrees Celsius under 5% carbon dioxide.
In a laminar flow hood under sterile conditions, renew the culture medium with 10 milliliters of warm Mbb 27 low medium.
Incubate for 2 days in a humidified incubator at 37 degrees Celsius under 5% carbon dioxide.
To harvest the OCM, collect the supernatant containing oligodendrocyte-secreted factors. Filter-sterilize the OCM using a 0.22-micron filter.
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