scholarly journals Real-Time Multidepth Multiphoton Microscopy Using Pixel-to-Pixel Focus-Switching

2020 ◽  
Vol 10 (20) ◽  
pp. 7173
Author(s):  
Yifan Qin ◽  
Deying Chen ◽  
Yuanqin Xia
Keyword(s):  
Real Time ◽  
Imaging System ◽  
Second Harmonic ◽  
Multiphoton Microscopy ◽  
Cost Effective ◽  
Axial Direction ◽  
Three Dimensions ◽  
Mouse Lung ◽  
Label Free

Multiphoton microscopy is a well-established technique for biomedical applications, but real-time multidepth multimodal multiphoton microscopy using non-imaging detection has barely been discussed. We demonstrate a novel label-free imaging system capable of generating multimodal multiphoton signals at different focal planes simultaneously. Two spatially overlapped and temporally interlaced beams are obtained by applying cost-effective electro-optic modulator (EOM)-based fast-switching light paths. The switching beams have different divergence properties, enabling imaging at different depths into samples. The EOM is synchronized to the pixel clock from the microscope, achieving pixel-to-pixel focus-switching. The capability of the imaging system is demonstrated by performing real-time multidepth two-photon fluorescence (TPF) and second-harmonic generation (SHG) imaging of freshly excised mouse lung lobes. TPF and SHG images are acquired at two wavelength ranges. One is between 415 and 455 nm, and the other is between 495 and 635 nm. The microenvironment of pulmonary alveoli is depicted by the distributions of both elastin fibers visualized by TPF and collagen fibers illustrated by SHG. Macrophages residing inside apparent alveolar lumens are also identified by TPF, which shows that the imaging system is capable of localizing biological objects in three dimensions and has the potential of monitoring in vivo cellular dynamics in the axial direction.

MULTIPHOTON MICROSCOPY: A NEW APPROACH, IN PHYSIOLOGICAL STUDIES AND PATHOLOGICAL DIAGNOSIS FOR OPHTHALMOLOGY

2009 ◽  
Vol 02 (01) ◽  
pp. 45-60 ◽  
Author(s):  
CHIU-MEI HSUEH ◽  
WEN LO ◽  
SUNG-JAN LIN ◽  
TSUNG-JEN WANG ◽  
FUNG-RUNG HU ◽  
...  
Keyword(s):  
Second Harmonic ◽  
Healing Process ◽  
Multiphoton Microscopy ◽  
In Vivo Studies ◽  
Wound Healing Process ◽  
Label Free ◽  
Multiphoton Imaging ◽  
Morphological Alterations ◽  
Corneal Scar

Multiphoton microscopy (MPM), with the advantages of improved penetration depth, decreased photo-damage, and optical sectioning capability, has become an indispensable tool for biomedical imaging. The combination of multiphoton fluorescence (MF) and second-harmonic generation (SHG) microscopy is particularly effective in imaging tissue structures of the ocular surface. This work is intended to be a review of advances that MPM has made in ophthalmic imaging. The MPM not only can be used for the label-free imaging of ocular structures, it can also be applied for investigating the morphological alterations in corneal pathologies, such as keratoconus, infected keratitis, and corneal scar. Furthermore, the corneal wound healing process after refractive surgical procedures such as conductive keratoplasty (CK) can also be studied with MPM. Finally, qualitative and quantitative SHG microscopy is effective for characterizing corneal thermal denaturation. With additional development, multiphoton imaging has the potential to be developed into an effective imaging technique for in vivo studies and clinical diagnosis in ophthalmology.

scholarly journals Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Dimitrios Kapsokalyvas ◽  
Rodrigo Rosas ◽  
Rob W. A. Janssen ◽  
Jo M. Vanoevelen ◽  
Miranda Nabben ◽  
...  
Keyword(s):  
Second Harmonic ◽  
Multiphoton Microscopy ◽  
Light Sheet ◽  
Three Dimensions ◽  
Fluorescence Signal ◽  
Label Free ◽  
Microscopy Imaging ◽  
Isotropic Resolution ◽  
Light Sheet Microscopy ◽  
3D Image Reconstruction

AbstractImaging in three dimensions is necessary for thick tissues and small organisms. This is possible with tomographic optical microscopy techniques such as confocal, multiphoton and light sheet microscopy. All these techniques suffer from anisotropic resolution and limited penetration depth. In the past, Multiview microscopy—imaging the sample from different angles followed by 3D image reconstruction—was developed to address this issue for light sheet microscopy based on fluorescence signal. In this study we applied this methodology to accomplish Multiview imaging with multiphoton microscopy based on fluorescence and additionally second harmonic signal from myosin and collagen. It was shown that isotropic resolution was achieved, the entirety of the sample was visualized, and interference artifacts were suppressed allowing clear visualization of collagen fibrils and myofibrils. This method can be applied to any scanning microscopy technique without microscope modifications. It can be used for imaging tissue and whole mount small organisms such as heart tissue, and zebrafish larva in 3D, label-free or stained, with at least threefold axial resolution improvement which can be significant for the accurate quantification of small 3D structures.

scholarly journals Identifying Two Common Types of Breast Benign Diseases Based on Multiphoton Microscopy

Scanning ◽  
2018 ◽  
Vol 2018 ◽  
pp. 1-6 ◽  
Author(s):  
Yan Wu ◽  
Yuxiang Lin ◽  
Yuane Lian ◽  
Peihua Lin ◽  
Shu Wang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Imaging System ◽  
Second Harmonic ◽  
Multiphoton Microscopy ◽  
Normal Breast ◽  
Benign Diseases ◽  
Cystic Fluid ◽  
Two Photon ◽  
Time Diagnosis

Multiphoton microscopy has attracted increasing attention and investigations in the field of breast cancer, based on two-photon excited fluorescence (TPEF) and second-harmonic generation (SHG). However, the incidence of breast benign diseases is about 5 to 10 times higher than breast cancer; up to 30% of women suffer from breast benign diseases and require treatment at some time in their lives. Thus, in this study, MPM was applied to image fibroadenoma and fibrocystic lesion, which are two of the most common breast benign diseases. The results show that MPM has the capability to identify the microstructure of lobule and stroma in normal breast tissue, the interaction of compressed ducts with surrounding collagen fiber in fibroadenoma, and the architecture of cysts filled with cystic fluid in fibrocystic disease. These findings indicate that, with integration of MPM into currently accepted clinical imaging system, it has the potential to make a real-time diagnosis of breast benign diseases in vivo, as well as breast cancer.

scholarly journals Performance of a novel protease-activated fluorescent imaging system for intraoperative detection of residual breast cancer during breast conserving surgery

2021 ◽  
Vol 187 (1) ◽  
pp. 145-153
Author(s):  
Conor R. Lanahan ◽  
Bridget N. Kelly ◽  
Michele A. Gadd ◽  
Michelle C. Specht ◽  
Carson L. Brown ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Real Time ◽  
Imaging System ◽  
Ex Vivo ◽  
Menopausal Status ◽  
Fluorescent Imaging ◽  
Cancer Subtypes ◽  
Surgical Workflow ◽  
Tumor Types

Abstract Purpose Safe breast cancer lumpectomies require microscopically clear margins. Real-time margin assessment options are limited, and 20–40% of lumpectomies have positive margins requiring re-excision. The LUM Imaging System previously showed excellent sensitivity and specificity for tumor detection during lumpectomy surgery. We explored its impact on surgical workflow and performance across patient and tumor types. Methods We performed IRB-approved, prospective, non-randomized studies in breast cancer lumpectomy procedures. The LUM Imaging System uses LUM015, a protease-activated fluorescent imaging agent that identifies residual tumor in the surgical cavity walls. Fluorescent cavity images were collected in real-time and analyzed using system software. Results Cavity and specimen images were obtained in 55 patients injected with LUM015 at 0.5 or 1.0 mg/kg and in 5 patients who did not receive LUM015. All tumor types were distinguished from normal tissue, with mean tumor:normal (T:N) signal ratios of 3.81–5.69. T:N ratios were 4.45 in non-dense and 4.00 in dense breasts (p = 0.59) and 3.52 in premenopausal and 4.59 in postmenopausal women (p = 0.19). Histopathology and tumor receptor testing were not affected by LUM015. Falsely positive readings were more likely when tumor was present < 2 mm from the adjacent specimen margin. LUM015 signal was stable in vivo at least 6.5 h post injection, and ex vivo at least 4 h post excision. Conclusions Intraoperative use of the LUM Imaging System detected all breast cancer subtypes with robust performance independent of menopausal status and breast density. There was no significant impact on histopathology or receptor evaluation.

scholarly journals Real-Time In Vivo Bioluminescent Imaging for Evaluating the Efficacy of Antibiotics in a Rat Staphylococcus aureus Endocarditis Model

2005 ◽  
Vol 49 (1) ◽  
pp. 380-387 ◽  
Author(s):  
Yan Q. Xiong ◽  
Julie Willard ◽  
Jagath L. Kadurugamuwa ◽  
Jun Yu ◽  
Kevin P. Francis ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Real Time ◽  
Imaging System ◽  
Bioluminescent Imaging ◽  
Vancomycin Therapy ◽  
Treatment Groups ◽  
Highly Sensitive ◽  
Relevant Animal Model ◽  
And Control

ABSTRACT Therapeutic options for invasive Staphylococcus aureus infections have become limited due to rising antimicrobial resistance, making relevant animal model testing of new candidate agents more crucial than ever. In the present studies, a rat model of aortic infective endocarditis (IE) caused by a bioluminescently engineered, biofilm-positive S. aureus strain was used to evaluate real-time antibiotic efficacy directly. This strain was vancomycin and cefazolin susceptible but gentamicin resistant. Bioluminescence was detected and quantified daily in antibiotic-treated and control animals with IE, using a highly sensitive in vivo imaging system (IVIS). Persistent and increasing cardiac bioluminescent signals (BLS) were observed in untreated animals. Three days of vancomycin therapy caused significant reductions in both cardiac BLS (>10-fold versus control) and S. aureus densities in cardiac vegetations (P < 0.005 versus control). However, 3 days after discontinuation of vancomycin therapy, a greater than threefold increase in cardiac BLS was observed, indicating relapsing IE (which was confirmed by quantitative culture). Cefazolin resulted in modest decreases in cardiac BLS and bacterial densities. These microbiologic and cardiac BLS differences during therapy correlated with a longer time-above-MIC for vancomycin (>12 h) than for cefazolin (∼4 h). Gentamicin caused neither a reduction in cardiac S. aureus densities nor a reduction in BLS. There were significant correlations between cardiac BLS and S. aureus densities in vegetations in all treatment groups. These data suggest that bioluminescent imaging provides a substantial advance in the real-time monitoring of the efficacy of therapy of invasive S. aureus infections in live animals.

Real-time histology-like video of blood flow concatenated with the original label-free SLAM video collected from in vivo rat mammary tissue

ASVIDE ◽  
2020 ◽  
Vol 7 ◽  
pp. 220-220
Author(s):  
Yi Sun ◽  
Sixian You ◽  
Xiaoxi Du ◽  
Z. George Liu ◽  
Eric J. Chaney ◽  
...  
Keyword(s):  
Blood Flow ◽  
Real Time ◽  
Mammary Tissue ◽  
Label Free ◽  
Original Label

scholarly journals Real-Time Optical Monitoring of Endotracheal Tube Displacement

Biosensors ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 174
Author(s):  
Ramzan Ullah ◽  
Karl Doerfer ◽  
Pawjai Khampang ◽  
Faraneh Fathi ◽  
Wenzhou Hong ◽  
...  
Keyword(s):  
Endotracheal Tube ◽  
Real Time ◽  
Near Infrared ◽  
Ex Vivo ◽  
Light Emitting Diode ◽  
Clinical Care ◽  
Cost Effective ◽  
Infrared Light ◽  
In Vivo Experiments

Proper ventilation of a patient with an endotracheal tube (ETT) requires proper placement of the ETT. We present a sensitive, noninvasive, operator-free, and cost-effective optical sensor, called Opt-ETT, for the real-time assessment of ETT placement and alerting of the clinical care team should the ETT become displaced. The Opt-ETT uses a side-firing optical fiber, a near-infrared light-emitting diode, two photodetectors with an integrated amplifier, an Arduino board, and a computer loaded with a custom LabVIEW program to monitor the position of the endotracheal tube inside the windpipe. The Opt-ETT generates a visual and audible warning if the tube moves over a distance set by the operator. Displacement prediction is made using a second-order polynomial fit to the voltages measured from each detector. The system is tested on ex vivo porcine tissues, and the accuracy is determined to be better than 1.0 mm. In vivo experiments with a pig are conducted to test the performance and usability of the system.

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