Courses

PhD course Flow Cytometry & Cell Sorting

We run one-to-two courses yearly. The course is designed to equip those having only limited or no prior experience of flow cytometry with a basic grounding in the principles of the technique and working knowledge of its practice with special emphasis on the application of flow cytometry to cell sorting.

 

PhD courses - overview

 

 


Organizer
Associate Professor Søren Hansen, Dept. of Cancer & Inflammation, University of Southern Denmark, Odense.
Duration
 4 days
ECTS points: 2.4
Number of participants: Max. 8
Teachers: Morten Løbner(ML), BD Bioscience, Denmark, Claus H. Nielsen (CHN), Copenhagen University Hospital, Oriane Cedile (OC), Odense University Hospital, Rikke Sick Andersen (RSA), Agnieszka Wlodarczyk (AW), Søren Hansen (SH) Lars Vitved (LV) and Ulla Melchior Hansen (UMH), University of Southern Denmark
Location: Winsløwparken 21, University of Southern Denmark, Odense
Application deadline: 2 weeks prior to course start 
Registration: Tina Ludvig-Nymark, Ph.D kontoret Sundhedsvidenskab, tludvig-nymark@health.sdu.dk

Courses 2017 - 2018:       

                                     12. march – 15. march 2019

                                        22. october – 25.oktober 2019

Example of program:
Day 1
09.00-09.15 Introduction

 

09.15-10.45 Flow cytometry 1 The basic principles

                    - The flow system
                    - Laser excitation
                    - Light scatter and fluorescence
                    - Multiple fluorochrome compensation
                    - Autofluorescence
                    - Data analysis
                    - Quality control

 

10.45-11.00 Coffee

 

11.00-12.30 Flow cytometry 2 The basic practicalities

                    - The cell preparation
                    - The parameters for investigation (membrane glycoproteins, surface lipids,
                      intracellular proteins, DNA, free Ca2+, oxidative metabolites etc.)
                    - Labelling with fluorochromes (incl. choosing the right marker-fluorochrome
                      combination)
                    - Data acquisition – the software
                    - Gating
                    - Positive and negative controls

 

12.30-13.15 Lunch


Practical Sessions Winsløwparken 21, 1st floor, Flow Cyto Lab

13-15-16.15 Exercise 1 The basics

                   - Starting up
                   - Instrument settings
                   - Calibration with standard beads
                   - Compensation
                   - Maintenance

 

                      Exercise 2 Identification of cell sub-populations in a blood leukocyte preparation     
                   
                   - Morphological discrimination
                   - Identification with the help of surface markers (CD3, CD19 etc.)


Day2.(Theoretical sessions)
09.00-10.00 Flow cytometry 3 Antibodies as probes

                   - The antibody-antigen interaction
                   - Monoclonal vs. polyclonal antibodies
                   - Direct vs. indirect labelling
                   - The biotin-streptavidin connection
                   - Extracellular vs. intracellular probing (fixation and permeabilisation)

10.00-10.15 Coffee

10.15-11.15 Flow cytometry 4 Other probes
                   
DNA staining
                   - Free calcium staining
                   - Staining for membrane depolarisation
                   - Staining for oxidative metabolites
                   - Staining for detection of cell division
                   - Staining for markers of apoptosis

11.15-12.15 Flow cytometry 5 Quantitative flow cytometry

                   - Quantitation cell sub-populations
                   - Quantitation of surface markers
                   - Quantitation of ligand-receptor interactions
                   -  Cell-cycle analysis

12.15-13.00 Lunch


Practical Sessions Winsløwprken 21, 1 st floor, Flow Cyto Lab

Exercise 3 Viability assessment

                   - Apoptosis vs. necrosis (PI + Annexin V)

Exercise 4 Assessment of cell proliferation

                   - CFSE approach

Day 3 (Theoretical sessions)
09.00-11.45 *Flow cytometry & Cell sorting

                   - Basic considerations
                   - Important Factors for Successful Cell Sorting
                               Quality of the cell preparation 
                               Cell density
                               Sorting rate v. viability
                               Purity versus yield
                               “Invisible” impurities
                               Sorting strategies
                   - Alternatives to pehnotype sorting - side population sorting, reporter genes
                   - Practicalities  The time factor
                   - Recovery – the harsh reality
                   * including 15 min. coffee break

 

11.45-12-15 Discussion of results - Exercises 3 and 4

 

12.15-13-00 Lunch


Practical Sessions Winsløwparken 21, 1 st floor, Flow Cyto Lab

 

13.00-15.00   Exercise 6 (Demonstration) Introduction to the cell sorter

                    














 

 

 

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