In a scientific tutorial in 2011, one of us (W.T.) covered the fluorescent proteins (FPs) available at the time. As a follow-up, we will focus on recent developments in the field, including new FPs, new lasers, and the advances made in detecting FPs using spectral flow cytometry. We will discuss considerations for practical use of FPs and make recommendations for choice of FPs in flow cytometry. We will analyze FP data using fluorescence compensation for conventional flow cytometry and unmixing algorithms for spectral flow cytometry. Attendees are encouraged to bring along their own data. Upon completion of this tutorial, the attendee will have an overview of the properties of commonly used FPs, the pertinent information to create optimal combinations of FPs, the knowledge to devise feasible detection strategies, and the skill to analyze FP data.
Course Details or Outline:
- Basics/Overview Timeline of major developments in FPs List of commonly used FPs Considerations for practical use of FPs Recommendations for choice of FPs in flow cytometry
- Methods Strategies for simultaneous detection of multiple FPs Matching lasers with FPs Conventional vs spectral flow cytometry of FPs Flow cytometer calibration using FP beads
- Data Analysis Fluorescence compensation in conventional flow cytometry Unmixing algorithms in spectral flow cytometry Display of properly compensated FP data