Label-free, non-contact, in vivo ophthalmic imaging using photoacoustic remote sensing microscopy

AbstractIn vivo, minimally invasive microscopy in deep cortical and sub-cortical regions of the mouse brain has been challenging. To address this challenge, we present an in vivo high numerical aperture optical coherence microscopy (OCM) approach that fully utilizes the water absorption window around 1700 nm, where ballistic attenuation in the brain is minimized. Key issues, including detector noise, excess light source noise, chromatic dispersion, and the resolution-speckle tradeoff, are analyzed and optimized. Imaging through a thinned-skull preparation that preserves intracranial space, we present volumetric imaging of cytoarchitecture and myeloarchitecture across the entire depth of the mouse neocortex, and some sub-cortical regions. In an Alzheimer’s disease model, we report that findings in superficial and deep cortical layers diverge, highlighting the importance of deep optical biopsy. Compared to other microscopic techniques, our 1700 nm OCM approach achieves a unique combination of intrinsic contrast, minimal invasiveness, and high resolution for deep brain imaging.

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Effect of valproic acid on functional bleb morphology in a rabbit model of minimally invasive surgery

British Journal of Ophthalmology ◽

10.1136/bjophthalmol-2020-318691 ◽

2021 ◽

pp. bjophthalmol-2020-318691

Author(s):

Zhu Li Yap ◽

Li-Fong Seet ◽

Stephanie WL Chu ◽

Li Zhen Toh ◽

Farah Ilyana Ibrahim ◽

...

Keyword(s):

Valproic Acid ◽

Cell Growth ◽

Minimally Invasive ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Rabbit Model ◽

Terminal Deoxynucleotidyl Transferase ◽

Label Free ◽

Collagen Fibres

AbstractPurposeTo determine the effect of valproic acid (VPA) on bleb morphology and scar characteristics in a rabbit model of minimally invasive glaucoma surgery (MIGS).MethodsNine New Zealand white rabbits were subjected to MIGS with intraoperative implantation of the PreserFlo MicroShunt. Rabbits were then administered with subconjunctival injections of phosphate buffered saline (PBS) (n=4) or with VPA (n=5). Bleb morphology was examined by slit-lamp biomicroscopy and in vivo confocal microscopy. Postoperative day 28 tissues were examined by immunohistochemical evaluation and label-free multiphoton microscopy to visualise the collagen matrix, by terminal deoxynucleotidyl transferase dUTP nick-end labelling assay and immunofluorescent labelling for Ki67 expression to detect apoptosis and cell growth, and by real-time quantitative PCR to measure Col1a1, Fn, and Smad6 transcript expression.ResultsVPA-treated blebs were detectable on day 28, while the PBS-treated blebs were not detectable by day 14. VPA-treated blebs were diffuse, extended posteriorly with near normal conjunctival vascularity and featured a combination of reticular/blurred stromal pattern with evidence of relatively large stromal cysts. Instead of the deposition of thick, disorganised collagen fibres characteristic of the PBS bleb, the VPA bleb contained conspicuously thinner collagen fibres which were associated with similarly thinner fibronectin fibres. In corroboration, Col1a1 and Fn mRNA expression was reduced in the VPA blebs, while increased Smad6 expression implicated the disruption of the transforming growth factor beta pathway. Apoptosis and cell growth profiles appeared similar with both treatments.ConclusionsThe results support the application of VPA to enhance bleb morphology associated with good bleb function in MIGS with no apparent cytotoxicity.

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Applications of Vibrational Spectroscopy for Analysis of Connective Tissues

Molecules ◽

10.3390/molecules26040922 ◽

2021 ◽

Vol 26 (4) ◽

pp. 922

Author(s):

William Querido ◽

Shital Kandel ◽

Nancy Pleshko

Keyword(s):

Raman Spectroscopy ◽

Vibrational Spectroscopy ◽

Near Infrared ◽

Ex Vivo ◽

Biological Tissues ◽

Label Free ◽

Connective Tissues ◽

Bone Properties ◽

Composition And Structure

Advances in vibrational spectroscopy have propelled new insights into the molecular composition and structure of biological tissues. In this review, we discuss common modalities and techniques of vibrational spectroscopy, and present key examples to illustrate how they have been applied to enrich the assessment of connective tissues. In particular, we focus on applications of Fourier transform infrared (FTIR), near infrared (NIR) and Raman spectroscopy to assess cartilage and bone properties. We present strengths and limitations of each approach and discuss how the combination of spectrometers with microscopes (hyperspectral imaging) and fiber optic probes have greatly advanced their biomedical applications. We show how these modalities may be used to evaluate virtually any type of sample (ex vivo, in situ or in vivo) and how “spectral fingerprints” can be interpreted to quantify outcomes related to tissue composition and quality. We highlight the unparalleled advantage of vibrational spectroscopy as a label-free and often nondestructive approach to assess properties of the extracellular matrix (ECM) associated with normal, developing, aging, pathological and treated tissues. We believe this review will assist readers not only in better understanding applications of FTIR, NIR and Raman spectroscopy, but also in implementing these approaches for their own research projects.

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Recovery of photoacoustic images based on accurate ultrasound positioning

Visual Computing for Industry Biomedicine and Art ◽

10.1186/s42492-021-00072-2 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Yinhao Pan ◽

Ningbo Chen ◽

Liangjian Liu ◽

Chengbo Liu ◽

Zhiqiang Xu ◽

...

Keyword(s):

Biological Tissue ◽

Large Field ◽

Photoacoustic Microscopy ◽

Label Free ◽

Imaging Data ◽

Motor Speed ◽

Image Correction ◽

Microscopy Imaging ◽

Scanning Method

AbstractPhotoacoustic microscopy is an in vivo imaging technology based on the photoacoustic effect. It is widely used in various biomedical studies because it can provide high-resolution images while being label-free, safe, and harmless to biological tissue. Polygon-scanning is an effective scanning method in photoacoustic microscopy that can realize fast imaging of biological tissue with a large field of view. However, in polygon-scanning, fluctuations of the rotating motor speed and the geometric error of the rotating mirror cause image distortions, which seriously affect the photoacoustic-microscopy imaging quality. To improve the image quality of photoacoustic microscopy using polygon-scanning, an image correction method is proposed based on accurate ultrasound positioning. In this method, the photoacoustic and ultrasound imaging data of the sample are simultaneously obtained, and the angle information of each mirror used in the polygon-scanning is extracted from the ultrasonic data to correct the photoacoustic images. Experimental results show that the proposed method can significantly reduce image distortions in photoacoustic microscopy, with the image dislocation offset decreasing from 24.774 to 10.365 μm.

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In Vitro and In Vivo Multispectral Photoacoustic Imaging for the Evaluation of Chromophore Concentration

Sensors ◽

10.3390/s21103366 ◽

2021 ◽

Vol 21 (10) ◽

pp. 3366

Author(s):

Aneline Dolet ◽

Rita Ammanouil ◽

Virginie Petrilli ◽

Cédric Richard ◽

Piero Tortoli ◽

...

Keyword(s):

Remote Sensing ◽

Photoacoustic Imaging ◽

Hyperspectral Remote Sensing ◽

Reference Spectrum ◽

In Vivo Experiments ◽

Mouse Tumors ◽

Chromophore Concentration ◽

Saturation Rate

Multispectral photoacoustic imaging is a powerful noninvasive medical imaging technique that provides access to functional information. In this study, a set of methods is proposed and validated, with experimental multispectral photoacoustic images used to estimate the concentration of chromophores. The unmixing techniques used in this paper consist of two steps: (1) automatic extraction of the reference spectrum of each pure chromophore; and (2) abundance calculation of each pure chromophore from the estimated reference spectra. The compared strategies bring positivity and sum-to-one constraints, from the hyperspectral remote sensing field to multispectral photoacoustic, to evaluate chromophore concentration. Particularly, the study extracts the endmembers and compares the algorithms from the hyperspectral remote sensing domain and a dedicated algorithm for segmentation of multispectral photoacoustic data to this end. First, these strategies are tested with dilution and mixing of chromophores on colored 4% agar phantom data. Then, some preliminary in vivo experiments are performed. These consist of estimations of the oxygen saturation rate (sO2) in mouse tumors. This article proposes then a proof-of-concept of the interest to bring hyperspectral remote sensing algorithms to multispectral photoacoustic imaging for the estimation of chromophore concentration.

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Subwavelength-resolution label-free photoacoustic microscopy of optical absorption in vivo

Optics Letters ◽

10.1364/ol.35.003195 ◽

2010 ◽

Vol 35 (19) ◽

pp. 3195 ◽

Cited By ~ 154

Author(s):

Chi Zhang ◽

Konstantin Maslov ◽

Lihong V. Wang

Keyword(s):

Optical Absorption ◽

Photoacoustic Microscopy ◽

Label Free ◽

Subwavelength Resolution

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Simulation and Analysis of Highly Sensitive MEMS Cantilever Designs for “in vivo Label Free” Biosensing

Procedia Technology ◽

10.1016/j.protcy.2014.08.012 ◽

2014 ◽

Vol 14 ◽

pp. 85-92 ◽

Cited By ~ 7

Author(s):

Deep Kishore Parsediya ◽

Jawar Singh ◽

Pavan Kumar Kankar

Keyword(s):

Label Free ◽

Highly Sensitive

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Distinct roles of tumor associated mutations in collective cell migration

Scientific Reports ◽

10.1038/s41598-021-89130-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Rachel M. Lee ◽

Michele I. Vitolo ◽

Wolfgang Losert ◽

Stuart S. Martin

Keyword(s):

Collective Behavior ◽

Metastatic Potential ◽

Breast Epithelial Cell ◽

Collective Cell Migration ◽

Label Free ◽

Collective Migration ◽

Contrast Imaging ◽

Pten Deletion

AbstractRecent evidence suggests that groups of cells are more likely to form clinically dangerous metastatic tumors, emphasizing the importance of understanding mechanisms underlying collective behavior. The emergent collective behavior of migrating cell sheets in vitro has been shown to be disrupted in tumorigenic cells but the connection between this behavior and in vivo tumorigenicity remains unclear. We use particle image velocimetry to measure a multidimensional migration phenotype for genetically defined human breast epithelial cell lines that range in their in vivo behavior from non-tumorigenic to aggressively metastatic. By using cells with controlled mutations, we show that PTEN deletion enhances collective migration, while Ras activation suppresses it, even when combined with PTEN deletion. These opposing effects on collective migration of two mutations that are frequently found in patient tumors could be exploited in the development of novel treatments for metastatic disease. Our methods are based on label-free phase contrast imaging, and thus could easily be applied to patient tumor cells. The short time scales of our approach do not require potentially selective growth, and thus in combination with label-free imaging would allow multidimensional collective migration phenotypes to be utilized in clinical assessments of metastatic potential.

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