The ability of silica nanoparticles conjugated to Bromelain to penetrate tumor ECM was analyzed by measuring the migration distance of fluorescent nanoparticles through a column of matrigel. Matrigel was pipeted into a 96 well plate and incubated. After gel formation, wells were treated with 100 μg of MSN dispersed in PBS, 100 μg of Br–MSN dispersed in PBS, and pure PBS as a control. The plate was incubated for 24 h and Z-stacks were obtained using a Nikon Eclipse Ti Inverted Fluorescent Microscope (Nikon Instruments, Inc., Melvillle, NY). The position of the fluorescent nanoparticle migration front was recorded and analyzed using NIS Elements AR software (Nikon Instruments, Inc.).
Eclipse ti inverted fluorescent microscope
Manufactured by Nikon
Sourced in Japan
The Eclipse Ti inverted fluorescent microscope is a high-performance laboratory equipment designed for advanced imaging applications. It features a robust and versatile design that enables researchers to conduct detailed fluorescence microscopy experiments. The core function of this microscope is to provide clear and accurate visualization of labeled samples, allowing for the observation and analysis of cellular structures, molecular interactions, and other biological phenomena.
Automatically generated - may contain errors
Lab products found in correlation
Radius cell migration kits, by Cell Biolabs (2 mentions)
Nis elements ar software, by Nikon (1 mentions)
Triton x 100, by Thermo Fisher Scientific (1 mentions)
Bovine serum albumin (bsa), by GE Healthcare (1 mentions)
Anti cd31 89c2, by Cell Signaling Technology (1 mentions)
C2 confocal microscope, by Nikon (1 mentions)
Fluorogel 2 containing 4 6 diamidino 2 phenylindole dapi, by Electron Microscopy Sciences (1 mentions)
Si8000 motion imaging system, by Sony (1 mentions)
Stage top incubator, by Tokai Hit (1 mentions)
Prolong gold antifade, by Thermo Fisher Scientific (1 mentions)
Nis elements c software, by Nikon (1 mentions)
Alexa fluor conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
H 3300, by Vector Laboratories (1 mentions)
Tcs sp5 confocal microscope, by Nikon (1 mentions)
Coolsnap120 es2 camera, by National Instruments (1 mentions)
Vectashield, by Vector Laboratories (1 mentions)
Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions)
Tissue tek oct compound, by Sakura Finetek (1 mentions)
Hematoxylin and eosin, by Merck Group (1 mentions)
Brilliant violet 605 anti mouse cd206, by BioLegend (1 mentions)
10 protocols using eclipse ti inverted fluorescent microscope
1
Assessing Nanoparticle Tumor Penetration
2
Immunofluorescence Staining of Endothelial Cells
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After 24, 48, or 72 h of incubation, samples were fixed in 4% paraformaldehyde (PFA), permeabilized with 0.1% Triton X-100 (Fisher Scientific, Waltham, MA, USA), blocked for 3 h with 4% bovine serum albumin (GE Healthcare Bio-Sciences, Chicago, IL, USA), incubated in primary (overnight) and secondary antibody (1 h), and mounted with FluoroGel II containing 4′,6-diamidino-2-phenylindole (DAPI) (Electron Microscopy Sciences, Hatfield, PA, USA) on glass slides. Primary antibodies used were anti-Glucose Transporter 1 (Glut-1) (ab15309, 1:200) (Abcam, Cambridge, MA, USA) and common endothelial markers such as anti-CD31 (89C2, 1:1000) (Cell Signaling Technology, Danvers, MA, USA) and VE-cadherin (2158, 1:100) (Cell Signaling Technology, Danvers, MA, USA) [35 (link),36 (link)]. With the immunostained samples, a Nikon Eclipse Ti inverted fluorescent microscope was used to acquire the wide-field epifluorescence images, while a Nikon C2 confocal microscope was used to acquire z-stacks of fluorescence images for 3-D reconstruction and analysis.
3
In situ Caspase-1 Activity Detection
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In situ detection of Caspase-1 activity was conducted as described previously (8 (link)). Briefly, unfixed eyes were enucleated and immediately placed in OCT mounting media and snap frozen in isopentane cooled by liquid nitrogen. Unfixed 5-μm-thick frozen sections of mouse eyes were incubated with CaspaLux1-E1D2 (Oncoimmunin) for 40 min at 37 °C in a humidified chamber. Afterward, slides were washed five times in PBS. Coverslips were placed on the tissue sections, and fluorescent and bright-field images were acquired on a Nikon Eclipse Ti inverted fluorescent microscope.
4
Electrical Stimulation of Myotubes
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Cells were differentiated on 6-well collagen gel plates prepared as described previously (refer “plate preparation”) and stimulated by EFS with C-Pace EP and 6-well C-Dish (IonOptics, Westwood, MA) for differentiated myotube maturation and muscle cell training under hypoxic conditions (5% O2, 5% CO2, 90% N2). The basic EFS stimulating protocol is described in Figure 2 . A movie was recorded and analyzed using an SI8000 Motion Imaging System (SONY, Tokyo, Japan) equipped with an Eclipse Ti inverted fluorescent microscope (Nikon) and a Stage Top Incubator (Tokai Hit, Shizuoka, Japan) to maintain cells under humidified conditions at 37°C and 5% CO2. Six-well hydrogel or collagen gel plates with cells were cultured in an incubator equipped with a C-Dish connected to the C-Pace EP. An EFS was applied at between 2-20V with a 2 ms interval and a 0.5-1 Hz. The movie was recorded for 270 frames at a frame rate of 27 f/s (equal to 10 s) and analyzed using associated software.
5
Cell Migration Assay with Radius Kit
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Cell migration assays were performed with Radius cell migration kits
(Cell Biolabs). MEFs (1×105 cells) were plated in 24-well
migration plates and allowed to attach overnight. The Radius gel was removed
according the manufacturer’s protocol. After gel removal, imaging was
performed over a 24h period with a Nikon Eclipse Ti inverted fluorescent
microscope and NIS elements (version 4.60) software. During image acquisition,
the environmental conditions were kept at 5% CO2 and 37°C
(Okolab). After the 24 hour period, cells were fixed and stained with DAPI
according to kit instructions. Cell migration was quantified by re-imaging
migration fields and counting DAPI-stained nuclei within the boundary of the
circular void left by the gel at 0h. Migration assays were performed in
triplicate.
(Cell Biolabs). MEFs (1×105 cells) were plated in 24-well
migration plates and allowed to attach overnight. The Radius gel was removed
according the manufacturer’s protocol. After gel removal, imaging was
performed over a 24h period with a Nikon Eclipse Ti inverted fluorescent
microscope and NIS elements (version 4.60) software. During image acquisition,
the environmental conditions were kept at 5% CO2 and 37°C
(Okolab). After the 24 hour period, cells were fixed and stained with DAPI
according to kit instructions. Cell migration was quantified by re-imaging
migration fields and counting DAPI-stained nuclei within the boundary of the
circular void left by the gel at 0h. Migration assays were performed in
triplicate.
6
Immunofluorescence Staining of Suspended Cells
7
Immunofluorescence Imaging of NF-κB Translocation
8
Subcellular Localization of TAK1 Variants
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U3013MG cells stably expressing C-terminally V5-tagged wild-type TAK1, 3xNLS::TAK1, or NES::TAK1 were seeded on PDL/laminin precoated glass cover slips and were stained two days after plating as follows: fixed in 4% PFA for 15 min, permeabilized and blocked with staining buffer (0.2% Triton X-100 and 2% goat serum in PBS) for 1 h, incubated with primary antibody in staining buffer overnight at 4 °C, next day washed three times in wash buffer (0.2% Triton X-100 in PBS), stained with secondary antibody in staining buffer for 1 h, washed three times in wash buffer followed by 5 min incubation with wash buffer containing 10 ng/ml DAPI and mounting on a glass slide with Vectashield (Vector). Images were acquired on an Eclipse Ti Inverted fluorescent microscope (Nikon) and processed using Fiji software (ImageJ).
9
Cell Migration Assay with Radius Kit
10
Immunofluorescence Staining of Mouse Ear Cryosections
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Explanted mice ears where cut by a biopsy pouch centered in the middle zone (diameter 8 mm), washed twice with PBS, embedded at the hedge in O.C.T. (Tissue-Tek O.C.T. Compound, Sakura Finetek), and instantly frozen in liquid nitrogen. 8-μm thick slides were obtained cutting ears block with a cryostat at −20 °C. For H&E staining, slides were thawed, hydrated, washed and stained with hematoxylin and eosin (Sigma-Aldrich). Immunofluorescence staining has been performed on consecutive ear sections. Briefly, slides were thawed and blocked with goat serum 5% (Sigma-Aldrich) PBS-T 1 × solution. After washing, they were incubated overnight at 4° with anti-macrophage antibody and anti- CD206 (Alexa Fluor 488 anti-mouse F4/80 and Brilliant Violet 605 anti-mouse CD206 Biolegend). Excess of the anti-neutrophil antibody was washed out with PBS 1X. Cells nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI). Slides were sealed with ProLong Gold antifade reagent (Life Technologies). Images were captured with a Nikon Eclipse Ti Inverted Fluorescent Microscope.
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