HepG2 cells were seeded at 25,000 cells/96 well. Following overnight incubation, the medium was changed to DMEM without supplements for an additional 24 h. Cells were subsequently incubated with or without PCSK9 (100 nM) and with or without PCSK9 inhibitors 5E11 (0.1–1.0 µM), evolocumab (0.1–1.0 µM) and suramin (200 µg/ml) in serum-free medium for 4 h at 37 °C before addition of 5 µg/ml DiI-LDL (Thermo Fisher Scientific) or BODIPY-LDL (Thermo Fisher Scientific) for 4 h at 37 °C. Wells were washed with PBS and cellular uptake of fluorescence was evaluated using an EnSpire Alpha Plate Reader (Perkin Elmer) at excitation/emission 552/573 nm (DiI) or 515/520 nm (BODIPY). After overnight incubation of plates at −80 °C, the number of cells/well was quantified using the CyQuant Cell Proliferation Assay (Thermo Fisher Scientific), according to the manufacturer’s protocol.
Bodipy
Manufactured by PerkinElmer
BODIPY is a fluorescent dye molecule that can be used as a labeling reagent in various biological and chemical applications. It has a characteristic blue-green emission spectrum and is known for its high fluorescence quantum yield and photostability. BODIPY can be attached to a wide range of biomolecules, such as proteins, nucleic acids, and lipids, to facilitate their detection and visualization.
Automatically generated - may contain errors
Lab products found in correlation
Cyquant cell proliferation assay, by Thermo Fisher Scientific (1 mentions)
Dii ldl, by Thermo Fisher Scientific (1 mentions)
Bodipy ldl, by Thermo Fisher Scientific (1 mentions)
Enspire alpha plate reader, by PerkinElmer (1 mentions)
Transwell insert, by Corning (1 mentions)
Bodipy, by Thermo Fisher Scientific (1 mentions)
Ultraview vox spinning disk, by PerkinElmer (1 mentions)
Fluorescence plate reader, by Agilent Technologies (1 mentions)
3 protocols using bodipy
1
PCSK9 Inhibitor Effects on LDL Uptake
2
Binding and Dissociation of Fluorescent LPS
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Binding and dissociation
of LPSs were performed using a previously published protocol.51 (link) Briefly, a stock solution of 100 μg/mL
of E. coli BODIPY-LPS [BODIPYFL (503/513),
Molecular Probes, Life Technologies, USA] was diluted to 10 μg/mL
in 1× phosphate-buffered saline (PBS; pH 7.4) and sonicated for
2 min every time prior to use. Two milliliters of this sonicated solution
of BODIPY-LPS was transferred to a quartz cuvette at a concentration
of 500 ng/mL. Subsequently, the respective concentrations of NCK-10
in PBS were added to it, and the fluorescence was measured using a
λS55 fluorescence spectrophotometer (PerkinElmer). The parameters
used were as follows: excitation wavelength: 485 nm; emission wavelengths:
500–700 nm; excitation slit width = 5 nm; emission slit width
= 15 nm; and temperature: 25 °C. Three independent experiments
were performed, and the data furnished are a representative image
of one experiment.
of LPSs were performed using a previously published protocol.51 (link) Briefly, a stock solution of 100 μg/mL
of E. coli BODIPY-LPS [BODIPYFL (503/513),
Molecular Probes, Life Technologies, USA] was diluted to 10 μg/mL
in 1× phosphate-buffered saline (PBS; pH 7.4) and sonicated for
2 min every time prior to use. Two milliliters of this sonicated solution
of BODIPY-LPS was transferred to a quartz cuvette at a concentration
of 500 ng/mL. Subsequently, the respective concentrations of NCK-10
in PBS were added to it, and the fluorescence was measured using a
λS55 fluorescence spectrophotometer (PerkinElmer). The parameters
used were as follows: excitation wavelength: 485 nm; emission wavelengths:
500–700 nm; excitation slit width = 5 nm; emission slit width
= 15 nm; and temperature: 25 °C. Three independent experiments
were performed, and the data furnished are a representative image
of one experiment.
3
Endothelial Fatty Acid Uptake Assay
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