Tightly packed monolayers will require some trituration to generate one-cell suspensions. Keep away from too much trituration or shear power by resting the pipette idea from the plate edge to keep up viability. If cells don't dissociate nicely, incubation time with dissociation reagent may need to be optimized.
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It can be crucial to quench the dissociation reagent using FACS buffer or quit medium, by adding a minimum of the identical or double the amount in the dissociation reagent.
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Notice: Progress to antibody staining if accomplishing Stay cell Evaluation soon after this phase. Alternatively, commence to fixable dye staining and PFA fixation.
Just in case cell clumping is noticed right after thawing cryopreserved cells, it is usually recommended to filter aggregated suspensions through 줄기세포 지방이식 a 37 µm cell strainer (e.g. Catalog #27250) for optimum results. In cases like this, the operate time on the CellPore™ Transfection System may possibly need to be enhanced to 10 seconds.
To evaluate the regional specificity of your little vs large airway, you could conduct the next assays:
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Sure, you’ll discover the action-by-move protocol for TEER measurement to evaluate the epithelial barrier integrity in ALI cultures right here.
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The next fluorochrome-conjugated antibodies and dyes are advised to facilitate analysis of gene-edited CD34+ HSPCs: