ER-Tracker Red, 20 μL

Product Information

Product Description/Introduction

ER-Tracker Red (Endoplasmic Reticulum Red Fluorescent Probe) is a combination of the red fluorescent dye BODIPY FL and Glibenclamide. The glibenclamide group in its chemical structure, originally a type II diabetes treatment drug, binds to the sulfonylurea receptor of ATP-sensitive potassium channels on the endoplasmic reticulum, and thus can be selectively localized to the endoplasmic reticulum and used as an endoplasmic reticulum-specific fluorescent probe for labeling the endoplasmic reticulum of cells.ER tracker red has good specificity, is highly selective for the endoplasmic reticulum of living cells, rarely stains mitochondria, and has low cytotoxicity. This probe can partially retain the staining characteristics of living cells after staining and fixation.

Storage and Shipping Conditions

Ship with dry ice; Store at -20 away from light for 12 months.

Product Content

Assay Protocol / Procedures

1. Assay working solution preparation:

1.1. This product is a 1 mM storage solution that is warmed back to room temperature prior to experimentation and centrifuged briefly at low speed to ensure that the reagents sink to the bottom of the tube;

1.2. It is diluted to 0.25-1 μM working solution for endoplasmic reticulum detection by buffer (Hanks, PBS, etc., Hanks is recommended) or basal medium (without serum);

2. Cell staining (the following protocol is for adherent cells; centrifugation is required for suspension cell exchange)

2.1. Normal or treated cells are washed 1-2 times with buffer for 3-5 min each time;

2.2. Add pre-warmed endoplasmic reticulum assay working solution, and incubate in a cell culture incubator at 37°C for 20 min (the incubation time can be adjusted to some extent due to different cell types and states);

2.3. Remove the endoplasmic reticulum assay working solution and wash with buffer 2-3 times for 3-5 min each;

2.4. Red fluorescence (Ex=587 nm, Em=615 nm) is observed under a fluorescence microscope after blocking the film directly or with an anti-fluorescence bursting agent (recommended G1401);

3. Cell fixation (optional):

Note: After staining, this product can be fixed, but the fluorescent signal will be attenuated after fixation; it cannot be perforated for processing, and perforation will lead to the disappearance of fluorescent labeling.

3.1. The stained and washed treated cells are added to the fixative (recommended G1101) and fixed for 5 min at room temperature;

3.2. Remove the fixative and wash with buffer 2-3 times for 3-5 min each time;

3.3. After washing, the cells can be subsequently blocked, microscopically examined or further stained according to experimental needs.

Note

1. Due to the different types and states of the cells, the working concentration of the dye, and the incubation time, can be adjusted appropriately.

2. The probe could not be perforated for treatment after staining, and cell fluorescence disappeared almost completely after perforation.

3. The pharmacologic properties of glibenclamide may affect certain functions of the endoplasmic reticulum; variable expression of sulfonylurea receptors in some specialized cells may also result in non-endoplasmic reticulum-specific staining.

4. When staining and washing live cells, the assay working solution and washing buffer need to be pre-warmed at 37°C. Staining and washing are accomplished by incubation in an incubator so as not to affect the morphology of the cells by temperature difference stimulation.

5. In order to prevent fluorescence quenching, the entire staining process needs to be operated away from light.

6. Probe master mixes should be stored in separate containers to avoid repeated freezing and thawing.To avoid waste, the staining solution is prepared as needed.

7. For your health and safety, please wear lab coat and gloves during operation.

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