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The following illustration is for preparing RNP complexes for 1 response. Modify appropriately depending on the volume of reactions necessary.
Get in touch with us to debate customized assay answers for many different other cell types to fit your certain needs.
Mucus could be washed from the floor on the cells when a week starting at 7 days 3, after the cells have been cultured in PneumaCult™-ALI Medium.
See details exhibiting excellent ALI culture morphology and better epithelial cell marker expression employing these suggested Transwell® inserts >
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Likely back for the plate, rinse Just about every perfectly with 1 mL of FACS buffer and transfer the volume for the 15 mL tube. Observe: Continue to keep cell suspension on ice after transfer towards the tube until finally willing to operate FACS.
Based on the level of mucus accumulation, a 2nd clean may be demanded. See how a mucus clean is executed in this ALI culture differentiation video (skip to 02:24) >
Sure, you’ll find the steps to perform an ICC staining on the epithelial cells cultured within the ALI With this protocol. Here's a summary of antibodies that may be used for the characterization of airway cultures:
We haven't identified any general performance distinctions involving coated and non-coated conditions; even so, collagen coating may perhaps enhance differentiation in a few donors, or if dealing with freshly isolated cells.
*This is a master resources record for the beneath protocols—all reagents talked about in Every single protocol are summarized.
Use cell counts to different the sample into two tubes – “unstained” and “with viability dye”.
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