The concentrations of MPO, MMPs and the cytokines IL-1β, IL-8, and tumor necrosis factor (TNF)-α in cell culture supernatants were measured using the DuoSet enzyme-linked immunosorbent assay (ELISA) development system (R&D Systems) and ELISA MAX Standard Set Human TNF-α (Cat# 430201; BioLegend, San Diego, CA, USA) according to manufacturer' instructions. The absorbance at 450 nm was measured on a microplate reader (2030 ARVO X4; PerkinElmer Japan; Tokyo).
2030 arvo x4
Manufactured by PerkinElmer
Sourced in Japan, United States
The 2030 ARVO X4 is a high-performance multimode microplate reader from PerkinElmer. It is designed to provide accurate and reliable detection of a wide range of assays, including fluorescence, luminescence, and absorbance-based measurements.
Automatically generated - may contain errors
Lab products found in correlation
Duoset enzyme linked immunosorbent assay, by R&D Systems (1 mentions)
Elisa development system, by R&D Systems (1 mentions)
Bz h4c, by Keyence (1 mentions)
Bz x800, by Keyence (1 mentions)
Celltiter 96 aqueous one solution cell proliferation assay, by Promega (1 mentions)
Calcein am, by PerkinElmer (1 mentions)
Tnf α, by Thermo Fisher Scientific (1 mentions)
Axio observer d1 microscope, by Zeiss (1 mentions)
Alamarblue cell viability reagent, by Thermo Fisher Scientific (1 mentions)
Capiject capillary blood collection tube, by Terumo (1 mentions)
6 protocols using 2030 arvo x4
1
Quantification of Inflammatory Markers
2
Quantifying Lipid Accumulation in Macrophages
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M0 and M2c macrophages were incubated with oxLDL at various concentrations for 24 h and stained with Oil Red O solution (ORO) (ScyTek Laboratories; Logan, UT, USA) as per manufacturer’s protocol. Cells were observed using a fluorescence microscope (model BZ-X800; Keyence; Osaka, Japan) equipped with 40× objective lens, and lipid droplet area was calculated using software program BZ-H4C (Keyence). Data were expressed as ratio of lipid droplet area to cell number in 10 randomly selected fields. Lipid quantity was determined using a microplate reader (2030 ARVO X4; PerkinElmer Japan; Tokyo) to measure absorbance at wavelength 490 nm of isopropanol-extracted supernatants from ORO-stained cells.
3
Fibroblast Cell Proliferation Assay
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Fibroblast sheets were soaked in 500 μL of a 1:4 mixture of (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) reagent (CellTiter 96 Aqueous One Solution Cell Proliferation Assay; Promega, Madison, WI, USA) and medium at 37 °C for 4 h. After incubation, 100 μL of the supernatant from each well was transferred to a 96-well plate and its absorbance was measured at 490 nm on a microplate reader (2030 ARVO X4; PerkinElmer, Boston, MA, USA).
4
Quantifying Tumor Cell Adhesion to Endothelial Cells
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Static adhesion assay using fluorescence-labeled tumor cells was performed. HUVECs (2.5×105 cells/well) pretreated with or without 10 ng/ml TNF-α (Invitrogen; Thermo Fisher Scientific) were cultured in 96-well plates overnight. TNF-α has the potential to stimulate endothelial cell adhesion. GIST-T1 and GIST-T1-CAD cells were labeled with 2 µg/ml Calcein-AM (Dōjindo Laboratories) at 37°C for 30 min, washed thrice with PBS, and resuspended at 2.5×106 cells/ml with serum-free DMEM, and followed by pipetting onto confluent HUVECs monolayers. After coculturing for 2 h, medium and unbounded tumor cells were removed and discarded. Adherent tumor cells and endothelial cells were washed three times with PBS. Then the amount of Calcein-AM fluorescence was measured using a fluorescence microplate reader (2030 ARVO X4, PerkinElmer Life and Analytical Sciences), at an excitation wavelength of 485 nm and emission wavelength of 530 nm.
5
Corneal Epithelial Cell Culture
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A research-grade cornea from a 69-year-old female (Sight Life) was processed and LECs were cultured with KGF and Y-27632 as described above, except that the concentration of KGF was varied among 0, 2.5, 5, 10, 20, 40, and 100 ng/ml. After 6 days of culture, cell viability was assessed using AlamarBlue Cell Viability Reagent (Thermo Fisher Scientific) before the cells reached confluency. The fluorescence intensity was measured using a microplate reader (2030 ARVO X4, PerkinElmer, Boston, MA, USA). Phase-contrast microscopic images were obtained using an Axio-observer D1 microscope (Carl Zeiss, Jena, Germany) at day 19.
6
Intestinal Permeability Assessment in Mice
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Mice administered with 2% DSS for 0, 3, and 6 days were fasted for 4 hours and gavaged with 4-kDa fluorescein isothiocyanate (FITC)-dextran (FD4, 600 mg/kg body weight, Sigma-Aldrich). Four hours after gavage, blood was then collected by cardiac puncture, and serum was separated by the use of Capiject Capillary Blood Collection Tubes (Terumo, Tokyo, Japan). The concentration of FITC-dextran in the serum was determined by measuring fluorescence at 535nm/485nm using spectrophotofluorometry (2030 ARVO X4, PerkinElmer, Waltham, MA) and then calculated according to a standard curve plotted by serially diluted FITC-dextran.
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