Tissues and biomaterials are composed of particles, including cells, nuclei, organelles, protein aggregates, lipid vesicles, and fibers. The size distribution of tissue particles and granularities is altered in various pathologies. These granularities span a wide range from nano- to micrometer scale, posing a unique challenge in quantifying the continuum of particles sizes in intact tissue. We introduce laser Speckle PARticle SizEr (SPARSE), a noncontact optical technique that enables particle sizing over 10 nanometer-to-10 micrometer range, through analyzing the spatiotemporal attributes of polarized laser speckle, back-scattered from biofluids and tissues. We demonstrate that SPARSE effectively quantifies the average particle sizes in milk, blood, and intact tissue without a prior knowledge of particles' refractive indices or concentrations. Through beam scanning, particle size distribution is mapped in benign and malignant breast tissues with high resolution (~100 micrometer), mirroring histopathological microstructures. By enabling particle sizing in intact biomaterials and tissues, SPARSE holds broad potential for applications across nanomedicine, diagnostics, and biotechnology.

Developing a functional tissue-engineered articular cartilage remains a challenge to improving clinical treatment of cartilage injury and joint-related degenerative disease. The dynamic self-regenerating cartilage (dSRC) approach presented here encourages autologous chondrocytes to generate their own matrix rather than imposing a matrix upon them. dSRC constructs were grown for 12 weeks under hypoxic conditions in reciprocating motion. Biochemical composition was evaluated, specifically water, collagen, and proteoglycan content. Speckle rHEologicAl micRoscopy (SHEAR) was utilized for spatially resolved evaluation of the shear modulus in engineered cartilage. Histological and immunohistochemical analyses of dSRC were also performed. The maturation of the dSRC matrix results in collagen and glycosaminoglycan (GAG) levels around 50% of those in native cartilage. SHEAR images demonstrate an increase in shear modulus of the matrix to ~20% that of native cartilage after 12 weeks. Histological support for excellent collagen and GAG production was evident, and immunohistochemistry showed a high preference for hyaline-like type II collagen in the neomatrix. A decrease in chondrocyte density occurred from an initial hypercellular matrix to that approaching native cartilage by 12 weeks. While this maturation of dSRC in vitro should not be construed as an absolute prediction of in vivo performance, these results are encouraging, representing a potential new cartilage repair and regeneration approach.

Fibrin plays an important role in both immune response and pathogen virulence during bacterial infection. Pathogens such as staphylococci interact with fibrin through dynamic processes, involving binding, entrapment, and release from fibrin scaffolds, through which they experience an evolving cascade of nano- to microscale dynamics that span broad timescales. Yet, the biophysical dynamics that unfold between invading bacteria and host fibrin are not well understood. Here, we present a non-invasive optical approach based on speckle fluctuation to characterize the multiscale dynamics of microparticles in fibrin scaffolds. Using microparticles of varying sizes and surface chemistry to emulate spherical, non-motile bacterial cells, we demonstrate real-time monitoring of bacteria-fibrin interactions during fibrin clot formation and fibrinolysis in purified fibrins scaffolds and clinical plasma clots. Our approach circumvents the need for sophisticated position tracking equipment, making it potentially applicable to a broad range of experimental systems for biophysical investigation of bacteria-extracellular network interactions.

Particle sizing of cellular structures may have significant diagnostic utility. We introduce a laser Speckle PARticle SizEr (SPARSE) that harnesses the spatio-temporal analysis of polarized speckle to estimate scattering particle sizes in opaque biofluids and tissues. SPARSE measurements significantly correlate with dynamic light scattering (DLS) in polybead suspensions (R2=0.91, p<.0001) and milk samples (R2=0.93, p<.0001). Similarly, in whole blood samples of increasing tonicities, SPARSE tracks RBC shrinkage in concordance with DLS (R2=0.84, p<.0001). Moreover, SPARSE maps of breast carcinoma reveal distinct sizes for adipose, fibrous, and epithelial compartments. These findings highlight the diagnostic potential of SPARSE in multiple conditions.

The viscoelastic properties and microstructure of the fibrin clot network are compromised in patients with coagulation abnormalities. In this study, we harness two novel optical approaches, iCoagLab and Spectrally Encoded Confocal Microscopy (SECM), to investigate how modifications in viscoelastic properties of blood clots are reflected in the microstructural features of the fibrin network in coagulopathic patients. We observe that blood clot viscoelasticity is directly related to fibrin density, fiber length, and fiber straightness of a fully polymerized fibrin network.

Alterations in tissue mechanical properties have emerged as not merely a symptom of cancer but a decisive regulator of various oncogenic processes. The tumor microenvironment exhibits a combination of elastic and viscous traits spanning a wide frequency spectrum, leading to a perceived paradox of both matrix stiffening and 'liquidization' that coexist during malignant transformation and metastasis. Yet, seminal mechanobiological studies have largely relied on a single mechanical descriptor of elasticity. We present wideband laser Speckle rHEologicAl micRoscopy (SHEAR) to investigate the elastic and viscous spectral signatures that exist over >5 decades of frequencies in various types of clinical tumor specimens.

Mechanical alterations that accompany neoplasia are believed to drive the hallmarks of cancer, including immune modulation. Here, we exploit laser Speckle rHEologicAl micRoscopy (SHEAR), to evaluate the shear viscoelastic modulus, G*(x,y,ω), of breast carcinoma specimens and to investigate the associations between G* and the presence of tumor-infiltrating lymphocytes(TILs). Results demonstrate that in high-TILs tumors, the spatially averaged |G*| is significantly reduced and the gradient of |G*| is significantly increased. Moreover, stroma-to-epithelium contrast of |G*| trends higher, whereas entropy trends lower. These outcomes suggest that the micro-mechanical properties of the tumor microenvironment likely correlate with the immune response.

Viscoelastic transformation of tissue drives aberrant cellular functions and is an early biomarker of disease pathogenesis. Tissues scale a range of viscoelastic moduli, from biofluids to bone. Moreover, viscoelastic behavior is governed by the frequency at which tissue is probed, yielding distinct viscous and elastic responses modulated over a wide frequency band. Existing tools do not quantify wideband viscoelastic spectra in tissues, leaving a vast knowledge gap. We present wideband laser speckle rheological microscopy (WB-SHEAR) that reveals elastic and viscous response over sub-megahertz frequencies previously not investigated in tissue. WB-SHEAR uses an optical, noncontact approach to quantify wideband viscoelastic spectra in specimens spanning a range of moduli from low-viscosity fibrin to highly elastic bone. Via laser scanning, micromechanical imaging is enabled to access wideband viscoelastic spectra in heterogeneous tumor specimens with high spatial resolution (25 micrometers). The ability to interrogate the viscoelastic landscape of diverse biospecimens could transform our understanding of mechanobiological processes in various diseases.