Sickle cell disease (SCD) exemplifies how molecular alterations cascade across biological scales to create complex pathophysiology. A single nucleotide substitution in the β-globin gene leads to polymerization of deoxygenated sickle hemoglobin (HbS), fundamentally altering red blood cell mechanics and blood flow dynamics. This chapter examines how quantitative biophysical and engineering methodologies have revolutionized our understanding of SCD across three interconnected scales: molecular HbS polymerization, cellular red blood cell mechanics, and tissue-level blood rheology. At the molecular scale, advanced techniques including electron microscopy, photolytic dissociation, and differential interference contrast microscopy have revealed the thermodynamics and kinetics of HbS fiber formation, establishing the double nucleation mechanism that explains both delayed onset and explosive polymer growth. Single-cell measurements using micropipette aspiration and microfluidic platforms have demonstrated the “all-or-nothing” mechanical transformation of red blood cells, with polymerization causing 100–1000-fold increases in cellular stiffness. At the tissue scale, bulk rheological studies and microfluidic approaches have identified pathological flow dynamics, including altered viscosity, yield stress behavior, and spatial flow inhomogeneities that concentrate rigid cells near vessel walls where they cause maximum endothelial damage. These multiscale insights have direct implications for improving clinical outcomes, from optimizing transfusion protocols to developing mechanism-based biomarkers for personalized treatment approaches. The integration of quantitative biophysical measurements across scales provides a powerful framework for understanding complex diseases and developing targeted interventions.