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Washable and compatible with serum-containing media after staining! ERseeing™ | Funakoshi

Date：July 27 2020Web Page No：81514

Funakoshi Co.,Ltd.

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.

ERseeing Labeling dye of ER

Limitations of Existing ER Stains vs. Advantages of ERseeing™

Comparison between ERseeing and conventional ER dye

The endoplasmic reticulum (ER) is a major organelle involved in various cellular processes. For fixed cells, ER can be visualized using immunofluorescence (IF) with antibodies targeting specific ER marker proteins (e.g., calnexin, calreticulin, PDI, BiP/GRP-78) or the ER retention signal KDEL. However, live-cell imaging is crucial for analyzing the physiological functions and dynamic morphology of the ER.

Conventional live-cell ER stains are typically fluorescently labeled K+ channel antagonists. Although widely used, they have significant limitations:

  • High Cytotoxicity/Artifacts: They block K+ channel activity, exerting a strong pharmacological impact on cell functions.
  • Reversible Binding: They wash out easily during medium exchange or washing steps, rapidly losing fluorescence signal.

ERseeing™ overcomes these challenges. It features a novel Rhodol-based green fluorescent dye conjugated to a protein-labeling group with a high affinity for the ER membrane composition. It selectively accumulates in the ER and irreversibly labels ER proteins. This allows users to replace the staining solution with serum-containing media, enabling long-term imaging under physiological conditions or drug-treated environments.


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Features

  • Standard Detection: Excitation / Emission: 509 nm / 524 nm. Compatible with standard FITC filter sets
  • Minimal Cell Disruption: Greatly reduces pharmacological side effects compared to traditional K+ channel blockers.
  • No-Wash Protocol Option: Cells can be imaged directly in the staining medium.
    Note: For maximum specificity and to avoid non-specific oil-droplet-like aggregations during extended incubation, washing before imaging is recommended.
  • Irreversible Retention: Permits post-staining medium exchange. Cells can be imaged in any preferred environment, including FBS-containing media.
  • Fixation Compatible: Allows cell fixation (e.g., PFA) after live-cell staining, enabling seamless integration with downstream immunofluorescence (IF) protocols.
    Note: This reagent is not suitable for staining pre-fixed cells; staining must be performed on live cells.

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Reference Data

Excitation and Emission Spectra

Compatible with standard green fluorescence (FITC) observation conditions.

Excitation and Emission Spectra of ERM
  • Excitation Spectrum: Blue / Emission Spectrum: Red (Peak Ex/Em: 509 nm / 524 nm).

Validation of ER Specificity

Co-staining with various organelle markers confirmed high ER specificity:

  • ER Markers: Exceptionally high co-localization (Pearson's correlation coefficient > 0.9).
  • Lysosome & Mitochondria Markers: No co-localization observed.
  • Golgi Marker: Partial co-localization observed. This indicates that some labeled proteins moved from the ER to the Golgi via the secretory pathway. (Co-localization with the Golgi decreases when an ER-to-Golgi transport inhibitor is added. For more detail, please see ☞ Reference.)
ER-Protein Capture Kit ER Specificity

Cellular Retention Evaluation

ERseeing Comparison of Retentivity

HeLa cells were stained with either ERseeing™ or a conventional ER stain for 15 minutes in serum-free medium. After replacing the solution with medium containing 10% FBS, the conventional stain showed a severe loss of fluorescence signal due to its reversible binding. In contrast, ERseeing™ retained a robust signal, proving its utility for flexible downstream imaging in serum-rich environments.

Low Cytotoxicity

Cytotoxicity Data (ERseeing)

CHO cells were treated with various concentrations of ERseeing™ for 18 hours, and cell viability was assessed via MTT assay. No toxicity was observed even at 3 μM, which is 30 times higher than the recommended working concentration of 100 nM.

Combined Use with Immunofluorescence (IF)

Combined Use with IF (ERseeing)

HeLa cells were stained with ERseeing™ (100 nM) while alive, fixed with 4% PFA, and then subjected to immunofluorescence using an anti-calnexin antibody. Thanks to irreversible protein labeling, ERseeing™ withstands the fixation and permeabilization processes required for multiplexed antibody staining.


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Users' Voice

Users_Voice

"Unlike conventional reagents, ERseeing™ helps us observe the ER clearly with high specificity, even in samples where ER function has deteriorated. It is an excellent reagent."

— Feedback from a university-based researcher


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Reference

  • Fujisawa, A., et al., "Chemical Profiling of the Endoplasmic Reticulum Proteome Using Designer Labeling Reagents.", J. Am. Chem. Soc., 140, 17060-17070 (2018). [PMID: 30433779]

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Citation

  • Miyasato, S., et al., "Constitutively active GPR43 is crucial for proper leukocyte differentiation", FASEB J., 37(1), e22676(2023). [PMID: 36468834]
  • Mori, S., et al., "Intracellular TAS2Rs act as a gatekeeper for the excretion of harmful substances via ABCB1 in keratinocytes", FASEB Bioadv., 6(10), 424-441(2024). [PMID: 39372126]

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Product Information

[Date : July 22 2026 00:11]

Detail Product Name Product Code Supplier Size Price
ERseeing, Endoplasmic Reticulum Green
DatasheetThis may not be the latest data sheet. 		FDV-0038 FNAFunakoshi Co.,Ltd. 10 nmol $380

Description ERseeing is irreversible ER labeling dye and suitable for long-term live imaging.
Storage -20°C CAS
Link

ERseeing™(Endoplasmic Reticulum Green) in a minute
ER-Protein Capture Kit | Funakoshi

[Date : July 22 2026 00:11]

ERseeing, Endoplasmic Reticulum Green

DatasheetThis may not be the latest data sheet.


  • Product Code: FDV-0038
  • Supplier: FNA
  • Size: 10nmol
  • Price: $380

Description ERseeing is irreversible ER labeling dye and suitable for long-term live imaging.
Storage -20°C CAS
Link

ERseeing™(Endoplasmic Reticulum Green) in a minute
ER-Protein Capture Kit | Funakoshi



Related Product

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

[Date : July 22 2026 00:11]

Detail Product Name Product Code Supplier Size Price
ER-Protein Capture Kit
DatasheetThis may not be the latest data sheet. 		FDV-0039 FNAFunakoshi Co.,Ltd. 1 kit $600

Description ER-Protein Capture Kit enables to purify ER associated proteins to evaluate various ER roles with easy procedures and no special equipment such as ultracentrifuge machine.
Storage -20°C CAS
Link

ER-Protein Capture Kit | Funakoshi

[Date : July 22 2026 00:11]

ER-Protein Capture Kit

DatasheetThis may not be the latest data sheet.


  • Product Code: FDV-0039
  • Supplier: FNA
  • Size: 1kit
  • Price: $600

Description ER-Protein Capture Kit enables to purify ER associated proteins to evaluate various ER roles with easy procedures and no special equipment such as ultracentrifuge machine.
Storage -20°C CAS
Link

ER-Protein Capture Kit | Funakoshi



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