ORGANELLE-ID-RGB^® reagent IV for microscopy

Manuals And Inserts

• Datasheet

Specific Protocol

Wide Field Fluorescence/Confocal Microscopy:

Reagent Preparation: Mix 2 μl of ORGANELLE-ID-RGB^® Reagent IV in 1 ml of buffer of choice. This volume is sufficient for 10 assays and may be scaled according to need.

Staining Adherent cells:

1. Grow cells directly onto glass slides or polystyrene tissue culture plates until ~80% confluent via standard tissue culture practices.
2. Remove growth media.
3. Dispense the freshly diluted staining solution in a volume sufficient for covering the cell monolayer.
4. Protect samples from light and incubate for 30 minutes at 37°C.
5. Remove the excess staining solution and, if necessary, add a few drops of buffer to prevent the cells from drying out.
6. Cover cells with a glass cover slip and observe under a fluorescence/confocal microscope with a filter set for DAPI (Ex/Em: 350/470nm), Texas Red (Ex/Em: 540/605 nm) and GFP/FITC (Ex/Em: 488/514 nm).

Staining Non-Adherent Cells:

1. Grow the cells via standard tissue culture practices.
2. Collect about 1 x 10⁵ cells. Centrifuge at 500 x g for 5 minutes. Remove supernatant.
3. Re-suspend cells in a volume of the freshly diluted staining solution sufficient for covering the cell pellet.
4. Protect the samples from light and incubate for 30 minutes at 37°C.
5. Centrifuge at 500 x g for 5 minutes. Remove supernatant.
6. Re-suspend the cells in 100 µl buffer.
7. Plate 10-15 µl of cells on a glass slide.
8. Cover cells with a glass cover slip and observe under a fluorescence/confocal microscope with a filter set for DAPI (Ex/Em: 350/470nm), Texas Red (Ex/Em: 540/605 nm) and GFP/FITC (Ex/Em: 488/514 nm).

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