Practical Course
Workshop/ Practical Course
HANDS-ON TECHNIQUES IN REGENERATIVE MEDICINE
for students, young doctors, and health specialists
Saturday 18 April 2026
Leonardo da Vinci Center of Competence in Personalized Medicine, 3D and Telemedicine, Robotic and Minimally Invasive Surgery, 5800 Pleven, Bulgaria
Venue:
Medical University Pleven,
PLEVEN, Bulgaria
PROGRAM WITH SYLLABI
WORKSHOP PROGRAM: MSC FUNCTIONALITY, ECM INTERACTIONS, NANOFIBERS DESIGN & 3D BIOPRINTING
1 Hands‑on: Mesenchymal Stromal Cells (MSC)
- Usual sources for obtaining MSCs
- MSC in culture:
- Short‑term and long‑term maintenance
- Passaging
- Freezing and storage
2 Functional Characterization of Adipose‑Derived MSCs
2.1 Adhesive Interaction with ECM Proteins: Collagen and Collagen Remodeling
(1 PPT slide)
2.2 Preparation of Collagen Type I – Native ECM Protein
(1 PPT slide: structure, affinity, receptors)
- Extraction of collagen from rat tails
- Printed protocol
- PPT slide
- Demonstration: tendon preparation, acetic acid extraction, desalting
- Fluorescent labeling of collagen with FITC
- Printed protocol
- PPT slide
- Demonstration of ready‑to‑use labeled collagen
- Preparation of collagen‑coated substrates
3 MSC Adhesion to Collagen
- Printed protocol
- PPT slide
- Harvesting MSCs: trypsin/EDTA, centrifugation, counting (all steps observed under phase contrast)
- Incubation of MSCs on collagen‑coated substrates (2 h, serum‑free)
-2 hour waiting period-
4 Hands-on Electrospinning of Nanofibers
- Overview of electrospinning equipment
- Short introduction to the method (PPT slide)
- Demonstration: electrospinning of collagen‑containing nanofibers
- Following printed protocol
- Demonstration of fiber alignment on rotating drum
- Phase‑contrast microscopy
- Mention only: nanofibers with secretome (PPT slide)
5 Hands‑on 3D Bioprinting
- Demonstration of bioprinting equipment
- PPT slide
- Advanced Biofabrication - overview by Murad Redzeb (2‑slide poster PPT)
- Demonstration of the method
6 MSC Adhesion on FITC‑Collagen (use already prepared samples from point 4)
- Observation under phase contrast
- FDA staining of living cells
- Demonstration on Thunder microscope
- Green fluorescence visualization
- Mention only: live‑dead assay (show image)
- Demonstration of immunofluorescent visualization:
- Actin cytoskeleton
- Focal adhesions
- Nucleus
- Comparison of MSC morphology on “good” vs. “bad” substrates
7 Remodeling of FITC‑Collagen by MSCs (1 PPT slide)
- Demonstration of FITC‑collagen substrates with adhering cells from point 4 and 7 (Thunder microscope)
- Explanation of remodeling process (since 5 h waiting is not possible)
-Show images demonstrating-:
- Dark zones of collagen removal
- Fibrillar reorganization
- Examples of good vs. poor remodeling
8 Motile Activity of MSCs on Collagen: Scratch Assay
- Demonstration of scratch creation with pipette tip
- Microscope adjustment on 3 scratches
- Start time‑lapse acquisition
- Show ready time‑lapse movies
- Explanation of cell migration parameter
- Mean migration speed - How to calculate
- Percentage of wound coverage- How to calculate