Golgi-Tracker Red

This plan only provides a guide, please modify it to meet your specific needs.

1. Prepare dyeing solution

(1) This product is provided in the form of 33.3mg/ml (~47mM) DMSO solution. It is recommended that when using it for the first time, the product should be concentrated at the bottom through low-speed centrifugation, and then packaged into small specifications according to the single dosage, and stored at -20℃ or -80℃ away from light and avoid repeated freezing and thawing.

(2) Prepare the working solution: Use a suitable buffer (such as PBS) to dilute the stock solution at a ratio of 1:100, and mix it to become the working solution.

Notice:

① Please adjust the concentration of the working fluid according to the actual situation and make it ready for use;

② The working concentration of Golgi-Tracker Red can be adjusted appropriately according to the actual situation. The recommended dilution ratio range is 1:50~1:200.

2. Cell suspension staining

(1) Suspension cells: Centrifuge at 1000g for 3-5 minutes, discard the supernatant, and wash twice with an appropriate amount of solution such as Hanks balanced salt solution (containing calcium and magnesium) or other buffers, for 5 minutes each time .

(2) Adherent cells: Wash twice with a suitable buffer, add trypsin to digest the cells, and centrifuge at 1000g for 3-5 minutes after digestion is completed.

(3) Use freshly prepared Golgi-Tracker Red staining working solution to resuspend the cells and incubate at 4°C in the dark for 30 minutes.

(4) Aspirate away the staining solution, wash the cells about 3 times with pre-cooled cell culture medium, replace with fresh culture medium, and incubate in the dark at 37°C for 30 minutes. ,

(5) Wash again with fresh culture medium, and then observe with a fluorescence microscope or confocal microscope.

3. Cell adhesion staining

(1) Culture adherent cells on sterile coverslips.

(2) Remove the coverslip from the culture medium, aspirate the excess culture medium, and place the coverslip in a humid environment.

(3) Remove the cell culture medium and use an appropriate amount of solution such as Hanks balanced salt solution (containing calcium and magnesium) or other buffers to wash the cells growing on the coverslip.

(4) Aspirate the rinse solution, add freshly prepared Golgi-Tracker Red dyeing working solution, and incubate at 4°C in the dark for 30 minutes.

(5) Aspirate away the staining solution, wash the cells about 3 times with pre-cooled cell culture medium, replace with fresh culture medium, and incubate in the dark at 37°C for 30 minutes.

(6) Wash again with fresh culture medium, and then observe with a fluorescence microscope or confocal microscope.

4. Microscopic examination: After the staining is completed, the Golgi apparatus shows bright and strong fluorescence staining, while other membrane systems in the cell show relatively weak fluorescence staining. The maximum excitation/emission light of Golgi-Tracker Red is 589/617 nm respectively.

Precautions:

1) Fluorescent dyes all have quenching problems. Please avoid light as much as possible to slow down fluorescence quenching.

2) For your safety and health, please wear a lab coat and disposable gloves.