This video features step-by-step instructions to help you plate perfect hepatocytes.
Sekisui XenoTech’s cryopreserved plateable hepatocytes are easy and convenient to use. Upon completing the thaw and resuspension process (https://www.xenotech.com/webinar-videos/cryopreserved-hepatocyte-thawing-demo-video), dilute the hepatocytes to the recommended cell concentration using OptiPlate Media.
Gently resuspend the hepatocyte suspension. You are now ready to seed the cells on the culture vessel. For culturing hepatocytes, the culture vessel must have a Collagen I coating.
Set your single channel pipet or repeator to the recommended seeding volume. Gently pipet the hepatocytes into each well, being sure to resuspend the cells periodically during the process.
Once the cells are dispensed, place the lid back on the culture vessel and transfer it to a 37°C humidified incubator. For even distribution of the cells, move the plate in a North/South East/West motion. The exception is the 96-well format. In this case, just place the plate in the incubator with no shaking.
After 2-4 hours of attachment, the cells are ready for media replacement. Remove the culture vessel from the incubator and return to the biosafety hood.
Sekisui XenoTech recommends a Matrigel Overlay for cells being cultured out to 5 days or longer. Matrigel is added to a final concentration of 0.25 mg/mL to ice cold OptiCulture media.
Mix the solution well.
Vigorously shake the plate to loosen unattached cells from each well. Tip the plate towards you to aspirate media from each well being careful not to touch the bottom of the well with aspirator tip.
Add overlay solution to each well, ensuring that media is being added towards the wall of each well as to not disrupt the attached cells.
Replace the lid on the culture vessel and return to incubator. Sekisui XenoTech recommends a 24 hour acclimation period.
After 24 hours you can proceed with feeding and dosing using Sekisui XenoTech’s OptiCulture media or begin long-term incubation experiments using Sekisui XenoTech’s OptIncubate media.
This concludes our hepatocyte plating demonstration video. If you have any questions or concerns, please contact Sekisui XenoTech at 913-438-7450 (toll-free in N. America at 1-877-588-7530), www.xenotech.com or firstname.lastname@example.org.