Significance: Current methods for analyzing pathological muscle tissue are time consuming and rarely quantitative, and they involve invasive biopsies. Faster and less invasive diagnosis of muscle disease may be achievable using marker-free in vivo optical sensing methods.

Aim: It was speculated that changes in the biochemical composition and structure of muscle associated with pathology could be measured quantitatively using visible wavelength optical spectroscopy techniques enabling automated classification.

Approach: A fiber-optic autofluorescence (AF) and diffuse reflectance (DR) spectroscopy device was manufactured. The device and data processing techniques based on principal component analysis were validated using in situ measurements on healthy skeletal and cardiac muscle. These methods were then applied to two mouse models of genetic muscle disease: a type 1 neurofibromatosis (NF1) limb-mesenchyme knockout (Nf1Prx1^{−  /  −}) and a muscular dystrophy mouse (mdx).

Results: Healthy skeletal and cardiac muscle specimens were separable using AF and DR with receiver operator curve areas (ROC-AUC) of >0.79. AF and DR analyses showed optically separable changes in Nf1Prx1^{−  /  −} quadriceps muscle (ROC-AUC >0.97) with no differences detected in the heart (ROC-AUC <0.67), which does not undergo gene deletion in this model. Changes in AF spectra in mdx muscle were seen between the 3 week and 10 week time points (ROC-AUC = 0.96) and were not seen in the wild-type controls (ROC-AUC = 0.58).

Conclusion: These findings support the utility of in vivo fiber-optic AF and DR spectroscopy for the assessment of muscle tissue. This report highlights that there is considerable scope to develop this marker-free optical technology for preclinical muscle research and for diagnostic assessment of clinical myopathies and dystrophies.

Significance: Changes in interstitial fluid clearance are implicated in many diseases. Using near-infrared (NIR) imaging with properly sized tracers could enhance our understanding of how venous and lymphatic drainage are involved in disease progression or enhance drug delivery strategies.

Aim: We investigated multichromatic NIR imaging with multiple tracers to assess in vivo microvascular clearance kinetics and pathways in different tissue spaces.

Approach: We used a chemically inert IR Dye 800CW (D800) to target venous capillaries and a purified conjugate of IR dye 680RD with 40 kDa PEG (P40D680) to target lymphatic capillaries in vivo. Optical imaging settings were validated and tuned in vitro using tissue phantoms. We investigated multichromatic NIR imaging’s utility in two in vivo tissue beds: the mouse tail and rat knee joint. We then tested the ability of the approach to detect interstitial fluid perturbations due to exercise.

Results: In an in vitro simulated tissue environment, free dye and PEG mixture allowed for simultaneous detection without interference. In the mouse tail, co-injected NIR tracers cleared from the interstitial space via distinct routes, suggestive of lymphatic and venous uptake mechanisms. In the rat knee, we determined that exercise after injection transiently increased lymphatic drainage as measured by lower normalized intensity immediately after exercise, whereas exercise pre-injection exhibited a transient delay in clearance from the joint.

Conclusions: NIR imaging enables simultaneous imaging of lymphatic and venous-mediated fluid clearance with great sensitivity and can be used to measure temporal changes in clearance rates and pathways.

Significance: Light-field fundus photography has the potential to be a new milestone in ophthalmology. Up-to-date publications show only unsatisfactory image quality, preventing the use of depth measurements. We show that good image quality and, consequently, reliable depth measurements are possible, and we investigate the current challenges of this novel technology.

Aim: We investigated whether light field (LF) imaging of the retina provides depth information, on which structures the depth is estimated, which illumination wavelength should be used, whether deeper layers are measurable, and what kinds of artifacts occur.

Approach: The technical setup, a mydriatic fundus camera with an LF imager, and depth estimation were validated by an eye model and in vivo measurements of three healthy subjects and three subjects with suspected glaucoma. Comparisons between subjects and the corresponding optical coherence tomography (OCT) measurements were used for verification of the depth estimation.

Results: This LF setup allowed for three-dimensional one-shot imaging and depth estimation of the optic disc with green light. In addition, a linear relationship was found between the depth estimates of the OCT and those of the setup developed here. This result is supported by the eye model study. Deeper layers were not measurable.

Conclusions: If image artifacts can be handled, LF technology has the potential to help diagnose and monitor glaucoma risk at an early stage through a rapid, cost-effective one-shot technology.

Significance: Selective plane illumination microscopy (SPIM) is an emerging fluorescent imaging technique suitable for noninvasive volumetric imaging of C. elegans. These promising microscopy systems, however, are scarce in academic and research institutions due to their high cost and technical complexities. Simple and low-cost solutions that enable conversion of commonplace wide-field microscopes to rapid SPIM platforms promote widespread adoption of SPIM by biologist for studying neuronal expressions of C. elegans.

Aim: We sought to develop a simple and low-cost optofluidic add-on device that enables rapid and immobilization-free volumetric SPIM imaging of C. elegans with conventional fluorescent microscopes.

Approach: A polydimethylsiloxane (PDMS)-based device with integrated optical and fluidic elements was developed as a low-cost and miniaturized SPIM add-on for the conventional wide-field microscope. The developed optofluidic chip contained an integrated PDMS cylindrical lens for on-chip generation of the light-sheet across a microchannel. Cross-sectional SPIM images of C. elegans were continuously acquired by the native objective of microscope as worms flowed in an L-shape microchannel and through the light sheet.

Results: On-chip SPIM imaging of C. elegans strains demonstrated possibility of visualizing the entire neuronal system in few seconds at single-neuron resolution, with high contrast and without worm immobilization. Volumetric visualization of neuronal system from the acquired cross-sectional two-dimensional images is also demonstrated, enabling the standard microscope to acquire three-dimensional fluorescent images of C. elegans. The full-width at half-maximum width of the point spread function was measured as 1.1 and 2.4  μm in the lateral and axial directions, respectively.

Conclusion: The developed low-cost optofluidic device is capable of continuous SPIM imaging of C. elegans model organism with a conventional fluorescent microscope, at high speed, and with single neuron resolution.