F-Actin staining of bovine foetal aorta endothelial cells using CytoPainter Phalloiding-iFluor 405 reagent

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CytoPainter Phalloidin-iFluor 405 Reagent was used to stain F-actin in Bovine Foetal Aeorta Endothelial (BFA) cells. This was tested on triton x-100 permeabilised and non-permeabilised cells, phalloidin was prepared at 1 in 1000, diluted in 1% BSA in PBS. Cells were stained for 20 minutes. Coverslips with cells were then prepared and used for confocal microscopy. Nucleus was also stained with a nuclear stain that is excited by 633 laser. Actin staining worked in both permeabilised and unpermeabilised cells, although fluorescence was dimmer when compared to phalloidin-488 or phalloidin-568 stain, however this could be overcome perhaps by leaving the product on the cells for longer. Fluoresence also bleached relatively quickly, so it was important not to excite the fluorophore too strongly or for too long. Despite this, cells stained well, showing the same actin structures seen with other phalloidin stains and had the advantage in that the product could be used more dilute (1 in 1000) compared to other stains that are used at 1 in 25 dilution. I would recommend this product if 488 or 568 fluorophores could not be used.

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Submitted Aug 14 2015

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