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 , 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 50 µm2 µm Array of 8 – 20 µm diameter wells Jurkat and HL60 cells trapped in 13.6 µm wells 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 Deflect cantilever until ~ 800pN applied or cell indented 3 µm HL60

HL60s stiffer than Jurkat cells and neutrophils consistent with a model of leukostasis Stiffness contributes to vessel blockage 855 Pa 48 Pa 156 Pa

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?