Invitrogen
Click-iT® EdU Alexa Fluor® 488 Imaging Kit
C10337
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I used the Click-iT® EdU Alexa Fluor® 488 Imaging Kit to perform proliferation assays of my iPSC derived mesenchymal cells and was very pleased with the results. The best advantage of this kit in my opinion is that the cells are not subjected to any harsh treatments hence not compromising the integrity and morphology of the cells. Therefore the protocol offers flexiblity to further co-stain with other markers which allowed me to perform cell type specific proliferation studies. The kit calls for multiple stock solutions to be prepared which is a limitation in terms for initial set-up of the assay. Other than that, I loved the kit.
Immunofluresence
iPSC-derived cells
The cells were incubated with 1 µM of EdU for 6 hours (user-defined). The cells were then fixed with 4% paraformaldehyde for 15 minutes at room temperature and washed with buffer containing 3% BSA. The cells were then permeablized with 0.5% triton X-100 for 20 minutes. The cells were then washed thoroughly with the same wash buffer and the Click-iT reaction cocktail (prepared in the same order as listed in the protocol) containing 1X Click-iT reaction buffer, copper sulphate, Alexa Fluor Azide and reaction buffer additive was added to the cells and incubated for 30 minutes at room temperature in the dark. The cells can then be washed and co-stained with other antibodies or DNA staining before imaging and analysis.
The Click-iT reaction cocktail has to be prepared in the same order as listed in the protocol.
The proliferating cells within my differentiated culture when co-stained with other cell-specific markers helped me identify the proliferating cell type.
None
Very bright and consistent staining
Initial set up of the kit is cumbersome
I would highly recommend it for proliferation assays
Product Name: Click-iT® EdU Alexa Fluor® 488 Imaging KitSupplier Page