Endoplasmic Reticulum (ER) staining dye for long-term live imaging ERseeing^™ | Funakoshi

ERseeing^™ is irreversible ER (Endoplasmic Reticulum) staining dye and suitable for long-term live imaging. Compared to conventional ER staining dye such as Glibenclamide-based dye, ERseeing^™ exhibits very low pharmacological effect and high retentivity which enables to visualize ER even after washout or medium change.

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Endoplasmic reticulum (ER) is the largest organelle in a cell and has unique and dynamic tubular or sheet structures. ER plays essential roles in protein synthesis and carbohydrate metabolism, and is a traffic origin of secreted pathway proteins including the Golgi apparatus, exocytosis, plasma membrane and extracellular proteins.

The most conventional ER dye is based on Glibenclamide fluorophore conjugate. Glibenclamide is known as a potent and specific inhibitor of the sulphonylurea receptors of ATP sensitive K+ channels. It selectively localized on ER, and then glibenclamide-based ER dyes can visualize ER structures.
However, this type of dye has problems as below.
- Negative pharmacological effect on K+ channel in ER.
- Washed out by wash step and medium change.

ERseeing^™ solves the above problems; it exhibits little effect on the ER functions pharmacologically, and enables to visualize ER even after washout or medium change. ERseeing^™ has two units, a rhodol type green fluorescent dye and a thioester type protein labeling group. Because this a rhodol type dye has a high affinity to ER membranes, right after adding the reagent to cells, it selectively and quickly accumulates into ER. Once it accumulated in ER, the labeling group of ERseeing^™ reacts with amino acids in ER-proteins randomly, and forms a covalent bond.

Comparison of ER specificity
HeLa cells were stained with ERseeing^™ (100 nM) and Glibencramide type ER staining. ERseeing^™ was highly overlapped with conventional Glibencramide type ER staining dye.

Comparison of retentivety
HeLa cells were treated with ERseeing^™ or Glibenclamide-based dye for 15 min and observed without wash (Left). Both reagents show ER staining. After that, cells were washed by PBS, added fresh media containing 10% FBS and observed again. While the Glibenclamide-based dye showed a very weak signal from the cells, ERseeing^™ maintains a good signal from ER. ERseeing^™ is suitable for long term imaging after medium change.

Excitation (blue) and fluorescent (red) spectrum.
Exmax /Emmax = 509/524 nm. Commercial FITC filter sets are compatible.

Endoplasmic Reticulum (ER) proteins purification / isolation kit : ER-Protein Capture Kit