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Can I use any of your Cell Meter™ Autophagy Assay Kits with a flow cytometer?

Posted June 12, 2019


If so, which one shows the best response? Can you provide a sample protocol? 

Answer

Yes, you can use any of our three Cell Meter™ Autophagy Assay Kits (Cat# 23000, 23001, 23002) with a flow cytometer, fluorescence microscope or fluorescence microplate reader. From our experience Cat# 23002 showed the best response when used in flow cytometric analysis, then Cat# 23001 and finally Cat# 23000.

Below is a sample autophagy assay protocol using a flow cytometer:

  1. Incubate Jurkat cells in RPMI medium with 10% FBS (control) or in 1X HBSS buffer with 5% FBS (starved) for 16 hours at 37°C.  
  2. Spin down both the control and starved cells, and then re-suspend in 1X HBSS buffer.
  3. Prepare dye working solution by diluting 20 μL of Autophagy Blue™, Autophagy Super Blue™, or Autophagy Green™ (depending on which kit you are using) with 10 mL of Stain Buffer.
  4. Stain cells with dye working solution at room temperature for 20-30 minutes.
  5. After staining, spin down cells and wash with 1X HBSS buffer.
  6. Analyze cells with flow cytometer.
    1. Autophagy Blue™ has fluorescence excitation and emission at Ex/Em = 335/520 nm
    2. Autophagy Super Blue™ has fluorescence excitation and emission at Ex/Em = 335/520 nm
    3. Autophagy Green™ has fluorescence excitation and emission at Ex/Em = 485/530 nm
Additional resources

Protocol for Cell Meter™ Autophagy Fluorescence Imaging Kit (Cat# 23002)

Cell Meter™ Autophagy Assay Kit *Blue Fluorescence*

Cell Meter™ Autophagy Fluorescence Imaging Kit

Cell Meter™ Autophagy Assay Kit *Green Fluorescence*