Transcript Force Microscopy of Non-adherent Cells: A Comparison of

Force Microscopy of Non-adherent Cells: A
Comparison of Leukemia Cell Deformability
Michael J. Rosenbluth, Wilbur A. Lam, and Daniel A. Fletcher
Biophysical Journal, 2006
Sophie Wong
20.309, MIT
November 20, 2008
Background
• AFM used to quantify mechanical properties
of biological material
• Current methods for measuring elasticity and
viscoelasticity
– Require indentation of cells adhered to substrate
– Not feasible for non-adherent cells
• Increased stiffness of lymphocytes may be the
cause of diabetes mellitus and leukemia
Three Goals
• Develop method for characterizing and
comparing deformability of leukemia cells
• Compare mechanical properties of
–myeloid (HL60) and lymphoid (Jurkat) lines
–normal neutrophils
• Compare Hertzian Mechanics Model vs.
Liquid Droplet Model
Developing the Method
• Used microfabricated wells to trap cells
• Array of 8 – 20 µm
diameter wells
•Jurkat and HL60 cells
trapped in 13.6 µm
wells
50 µm
2 µm
•Neutrophils trapped
in 10.8 µm wells
Deflection-position curve
• Cantilever deflection small compared to
indentation
• Piezo platform extended
at constant rate =
1506 nm/s
HL60
• Deflect cantilever until
~ 800pN applied or cell
indented 3 µm
HL60s stiffer than Jurkat cells and neutrophils
• consistent with a
model of
leukostasis
855 Pa
48 Pa
156 Pa
• Stiffness
contributes to
vessel blockage
Fitting the model
• Models used to determine cell elasticity and
viscoelasticity
• Hertzian Mechanics Model assumptions
– Homogeneous, Isotropic, Linear, Elastic (HILE)
– Material undergoes infinitesimally small strains
• Liquid Droplet Model assumptions
– Internal contents are homogeneous viscous
– Cortical tension constant around cell
– Cortical shell deforms around tip during indentation
– Radius of cell remains constant during indentation
Hertzian Model fits better
• Gray line = data
• Dash line = Hertzian mechanics model
• Dotted line = Liquid droplet model
Significance & Future Studies
• Deformability of leukemic cells plays
important role in leukostasis
• Where does difference in cell stiffness
originate?
– Filament networks?
– Cytoplasm?
– Nuclear or cell membranes?
• Investigate other factors involved in
leukostasis: adhesion, transmigration
Questions?