Design of twisted NIR BOPYIN with Highly Fluorescent Intensity for Monitoring Viscosity

Heat shock inductors have potential as treatment for degenerative and protein misfolding diseases. Dimethyl-sulfoxide is widely used as a solvent in pharmacological screening tests and has been shown to have heat shock induction effects. Transgenic Tg (hsp70l:EGFP-HRAS_G12V)io3(AB) zebrafish larvae were exposed for 24 hours to dimethyl-sulfoxide in concentratios of 0.1-2%, and to moderate heat shock inductors pentoxifylline and tacrolimus. Positive controls were exposed to 35, 38 and 40�C for 20 min, and incubated for 24 h at 28�C. Heat shock response was measured by fluorescence microscopy and signal intensity quantification in FIJI. Dimethyl-sulfoxide caused a dose-dependant increase in fluorescent intensity, but significantly lower compared with exposure to 38 and 40�C. Pentoxifylline and tacrolimus induced a significantly higher increase in fluorescence compared with 0.5% dimethyl-sulfoxide. Thus, although dimethyl-sulfoxide has independent heat shock induction effects, concentrations of up to 0.5% are suitable for heat shock response screening tests.

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Luminex Multiplex Bead Assay Monitoring HLA IgG Antibodies in Sensitized Pre- and Post-transplant Patients: Clonality of the Detection Antibody Impacts Specificity and Sensitivity

Applied Sciences ◽

10.3390/app11146430 ◽

2021 ◽

Vol 11 (14) ◽

pp. 6430

Author(s):

Mepur H. Ravindranath ◽

Narendranath M. Ravindranath ◽

Carly J. Amato-Menker

Keyword(s):

Igg Antibodies ◽

Fluorescent Intensity ◽

Heavy Chains ◽

Transplant Patients ◽

Specificity And Sensitivity ◽

Single Antigen ◽

Two Factors ◽

Specific Igg ◽

End Stage ◽

Multiplex Bead

The number and the binding affinity, measured as the mean fluorescent intensity (MFI) of HLA-specific IgG antibodies, formed in the sera of end-stage organ disease patients and allograft recipients, referred to as sensitization, may restrict the availability of a donor organ and/or lead to graft failure after transplantation. The MFI of HLA Abs in sera is monitored with the Luminex-based single-antigen bead (SAB) immunoassay. The following two factors may impact the reliable measurement of MFI: one, the HLA structural variants on the SAB, namely, trimeric HLA (closed conformers, CC) and monomeric heavy chains (open conformers, OC); and two, the nature of the detection Abs, namely, IgG heavy-chain binding polyclonal-Fab (IgHPolyFab) or Fc-binding monoclonal-IgG (FcMonoIgG). Anti-CC Abs correlate with positive flow cross-matches, and are considered to be pathogenic and damaging to the graft, whereas anti-OC Abs appear to have little relevance to graft attrition. The presence of both CC and OC on beads may impair the reliability of monitoring the nature and MFI of pathogenic Abs. Our objective is to compare the MFI of the HLA Abs in the sera of 20 sensitized patients in two different SAB assays, with the two detection Abs. Our data reveal that the admixture of OC with CC on beads will affect the reliability of the measurement of the pathogenic Abs, and that FcMonoIgG is the more sensitive and specific detection Ab for the accurate assessment of HLA sensitization.

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Automated method for the isolation of collecting ducts

AJP Renal Physiology ◽

10.1152/ajprenal.00273.2005 ◽

2006 ◽

Vol 291 (1) ◽

pp. F236-F245 ◽

Cited By ~ 18

Author(s):

R. Lance Miller ◽

Ping Zhang ◽

Tong Chen ◽

Andreas Rohrwasser ◽

Raoul D. Nelson

Keyword(s):

Flow Cytometry ◽

Large Particle ◽

Fluorescent Protein ◽

Collecting Duct ◽

Complex Object ◽

Cortical Collecting Duct ◽

Fluorescent Intensity ◽

Automated Method ◽

Collecting Ducts ◽

Time Required

The structural and functional heterogeneity of the collecting duct present a tremendous experimental challenge requiring manual microdissection, which is time-consuming, labor intensive, and not amenable to high throughput. To overcome these limitations, we developed a novel approach combining the use of transgenic mice expressing green fluorescent protein (GFP) in the collecting duct with large-particle-based flow cytometry to isolate pure populations of tubular fragments from the whole collecting duct (CD), or inner medullary (IMCD), outer medullary (OMCD), or connecting segment/cortical collecting duct (CNT/CCD). Kidneys were enzymatically dispersed into tubular fragments and sorted based on tubular length and GFP intensity using large-particle-based flow cytometry or a complex object parametric analyzer and sorter (COPAS). A LIVE/DEAD assay demonstrates that the tubules were >90% viable. Tubules were collected as a function of fluorescent intensity and analyzed by epifluorescence and phase microscopy for count accuracy, GFP positivity, average tubule length, and time required to collect 100 tubules. Similarly, mRNA and protein from sorted tubules were analyzed for expression of tubule segment-specific genes using quantitative real-time RT-PCR and immunoblotting. The purity and yield of sorted tubules were related to sort stringency. Four to six replicates of 100 collecting ducts (9.68 ± 0.44–14.5 ± 0.66 cm or 9.2 ± 0.7 mg tubular protein) were routinely obtained from a single mouse in under 1 h. In conclusion, large-particle-based flow cytometry is fast, reproducible, and generates sufficient amounts of highly pure and viable collecting ducts from single or replicate animals for gene expression and proteomic analysis.

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Abstract 3054: Comparison of Transforming Growth Factor-β1 Expression in Aortic Wall of Thoracic Aortic Aneurysm Versus Dissection

Circulation ◽

10.1161/circ.116.suppl_16.ii_684-b ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Shouguo Yang ◽

Guanggen Cui ◽

Ramin Beygui ◽

Fardad Esmailian ◽

Abbas Ardehali ◽

...

Keyword(s):

Aortic Aneurysm ◽

Thoracic Aortic Aneurysm ◽

Aortic Wall ◽

Transforming Growth Factor ◽

Smooth Muscle Actin ◽

Underlying Mechanism ◽

Transforming Growth Factor Β1 ◽

Fluorescent Intensity ◽

Nucleus Density ◽

Tgf Β1

Background The underlying mechanism of thoracic aortic aneurysm (TAA) and dissection(TAD) was undetermined, and one controversy lies in whether they represent the different dvelopement period of the same disorder or totally diferent diseases. This study is in aim to compare the expression and distribution of Transforming Growth Factors(TGF) β1 in the aortic wall of TAA versus TAD patients. Method Aortic specimens were obtained from patients underwent to aortic procedures for TAA (n=38) and TAD (n=20) at UCLA , and control aorta (CN) from organ donnor (n=20). Double immunofluorescent stainning of TGF-β1 and α-smooth muscle actin were performed with paraffin embeded slides for all aortic samples and semiquantified by fluorescent intensity analysis. Histopathologic examination were performed with HE, Verhoeff van-Gieson and Masson’s trichrome stain. Results TAA and TAD patients exhibited an up-regulation of TGF-β1 to 120.3% and 109.6% compared with CN separately (P<0.05), with TAA higher than TAD (P<0.05). TGF-β1 distributed unevenly across aortic wall with the highest levels expression in tunica media, followed by intima then adventitia. In intima, TGF-β1 was expressed at the same level for TAD as CN, but was increased to 115.2% for TAA compared to CN (P<0.05). In media, TGF-β1 increased by 127.2% in TAA and 116.1% in TAD compared to CN (P<0.01), with TAA being higher than TAD (P<0.05). In adventitia, TGF- β1 was up-regulated to 119.6% and 116.7% for TAA and TAD compared to CN (P<0.05). Nucleus density analysis showed cellular plasia in adventitia of TAA and TAD than CN (P<0.05 ), while TAD patients demonstrated a higher nucleus density than TAA in intima and adventitia (P<0.05). α-actin was increased in media of TAA and TAD to 164.5% and 120% than CN (P<0.01 and P<0.05). Attenuated and interrupted elastin and mild to severe cystic medial degeneration were characteristic histopathologic finding in 29 (76.3%) TAA and 17(85%) TAD patients. Conclusions TGF- β1 expression was up-regulated in aortic wall of TAA and TAD compared to CN. The significant higher levels of TGF- β1 in intima and media in TAA versus TAD patients implicated a probable positive effect of TGF- β1 to maintain aortic wall integrity, and/or greater comsamption of TGF- β1 in the aortic dissection.

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A Reduction Active Theranostic Nanoparticle for Enhanced Near-Infrared Imaging and Phototherapy by Reducing Glutathione Level in Cancer Cells

Journal of Nanoscience and Nanotechnology ◽

10.1166/jnn.2021.19514 ◽

2021 ◽

Vol 21 (12) ◽

pp. 5965-5971

Author(s):

Xiaofang Song ◽

Lifo Ruan ◽

Tianyu Zheng ◽

Jun Wei ◽

Jiayu Zhang ◽

...

Keyword(s):

Cancer Cells ◽

Disulfide Bonds ◽

Near Infrared ◽

Infrared Imaging ◽

Fluorescent Imaging ◽

Self Monitoring ◽

Fluorescent Intensity ◽

Turn On ◽

Hyperbranched Poly ◽

One Step

Facile preparation of a tumoral-stimuli-activated theranostic nanoparticle with simple constituents remains a challenge for tumor theranostic nanosystems. Herein we design a simple reductionresponsive turn-on theranostic nanoparticle for achieving fluorescent imaging and phototherapy combination. The theranostic nanoparticle is prepared by a simple one-step dialysis method of reduction active amphiphilic hyperbranched poly(β-amidoamines) and a near-infrared (NIR) dye indocyanine green (ICG). The fluorescence of ICG is quenched by the aggregation-caused quenching (ACQ) effect. The fluorescent intensity of free ICG at 816 nm was ∼40 times as high as that of particulate ICG. After reductive nanoparticles incubated with dithiothreitol (DTT), the size of the nanoparticles increased from 160 nm to 610 nm by Dynamic light scattering (DLS). As nanoparticles were internalized by cancer cells, the disulfide bonds would be cleaved by intracellular reduction agents like glutathione (GSH), leading to the release of entrapped ICG. The released ICG regained its fluorescence for self-monitoring the release and therapeutic effect of ICG by fluorescence spectra and the quantitative evaluation of NIR fluorescence intensity. Remarkably, nanoparticles can also reinforce antitumor efficacy through photodynamic therapy and GSH depletion property. This study provides new insights into designing turn-on theranostic systems.

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Perovskite Nanocrystals with Tunable Fluorescent Intensity during Anion Exchange for Dynamic Optical Encryption

ACS Applied Materials & Interfaces ◽

10.1021/acsami.1c14071 ◽

2021 ◽

Vol 13 (39) ◽

pp. 47072-47080

Author(s):

Yang Liu ◽

Yong Zhang

Keyword(s):

Anion Exchange ◽

Fluorescent Intensity ◽

Optical Encryption ◽

Perovskite Nanocrystals

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Method to minimize ozone effect on Cy5 fluorescent intensity in DNA microarrays

Analytical Biochemistry ◽

10.1016/j.ab.2017.09.001 ◽

2017 ◽

Vol 538 ◽

pp. 1-4

Author(s):

Youngjun Kim ◽

Hyun Hee Seo ◽

Mi Seon Jeong ◽

Ki Heon Lee ◽

In Ho Lee ◽

...

Keyword(s):

Dna Microarrays ◽

Fluorescent Intensity

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Time-resolved and temperature tuneable measurements of fluorescent intensity using a smartphone fluorimeter

The Analyst ◽

10.1039/c7an00535k ◽

2017 ◽

Vol 142 (11) ◽

pp. 1953-1961 ◽

Cited By ~ 13

Author(s):

Md Arafat Hossain ◽

John Canning ◽

Zhikang Yu ◽

Sandra Ast ◽

Peter J. Rutledge ◽

...

Keyword(s):

Steady State ◽

Fluorescence Intensity ◽

Time Resolved Fluorescence ◽

Time Resolved ◽

Fluorescent Intensity ◽

Intensity Measurements

A smartphone fluorimeter is demonstrated for steady-state and time-resolved fluorescence intensity measurements at tunable temperatures.

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Machine Learning Improves the Precision and Robustness of High-Content Screens

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057111414878 ◽

2011 ◽

Vol 16 (9) ◽

pp. 1059-1067 ◽

Cited By ~ 44

Author(s):

Peter Horvath ◽

Thomas Wild ◽

Ulrike Kutay ◽

Gabor Csucs

Keyword(s):

Machine Learning ◽

User Interaction ◽

Texture Features ◽

Large Data ◽

Complete Analysis ◽

Data Sets ◽

Reporter Protein ◽

Fluorescent Intensity ◽

Analysis Workflow

Imaging-based high-content screens often rely on single cell-based evaluation of phenotypes in large data sets of microscopic images. Traditionally, these screens are analyzed by extracting a few image-related parameters and use their ratios (linear single or multiparametric separation) to classify the cells into various phenotypic classes. In this study, the authors show how machine learning–based classification of individual cells outperforms those classical ratio-based techniques. Using fluorescent intensity and morphological and texture features, they evaluated how the performance of data analysis increases with increasing feature numbers. Their findings are based on a case study involving an siRNA screen monitoring nucleoplasmic and nucleolar accumulation of a fluorescently tagged reporter protein. For the analysis, they developed a complete analysis workflow incorporating image segmentation, feature extraction, cell classification, hit detection, and visualization of the results. For the classification task, the authors have established a new graphical framework, the Advanced Cell Classifier, which provides a very accurate high-content screen analysis with minimal user interaction, offering access to a variety of advanced machine learning methods.

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