CYTO-ID® Autophagy detection kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Incubate with Cyto-ID Green Detection Reagent for 30 min at 37ºC in the dark	Incase of Low CYTO-ID dye staining in all treatments, increase the reagent concentration (500X dilution of the dye is recommended) and the incubation time. Incase of High CYTO-ID staining, it could be that the cell culture medium was depleted of nutrients and so change media 4-8 hours before the experiment

Protocol tips
Incubate with Cyto-ID Green Detection Reagent for 30 min at 37ºC in the dark
Downstream tips
Incase of Low CYTO-ID dye staining in all treatments, increase the reagent concentration (500X dilution of the dye is recommended) and the incubation time. Incase of High CYTO-ID staining, it could be that the cell culture medium was depleted of nutrients and so change media 4-8 hours before the experiment

Publication protocol

Cyto-ID staining kit (Enzo Life Sciences, Farmingdale, NY) was used to detect autophagy in HEK293, Nthy-Ori-3 and TPC1 cells according to the manufacturer’s protocol. Briefly, cells were plated on cover slips at 70% confluency and allowed to attach overnight. Cell medium was aspirated followed by carefully washing twice using 1x Assay Buffer with 5% FBS. Cells were incubated with Cyto-ID Green Detection Reagent for 30 min at 37ºC in the dark. Cells were carefully washed twice with 1X Assay Buffer to remove excess dye and observed immediately under a confocal microscope.

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Manufacturer protocol

Download the product protocol from Enzo Life Sciences for CYTO-ID® Autophagy detection kit below.

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