Cultured human NP cells were rinsed with PBS three times, fixed with 4% formaldehyde for 15 min, and permeabilized with 0.3% Triton-X100 (Sigma, CO, USA) for 10 min. Subsequently, the cells were blocked with 6% BSA for 1 h and then were incubated overnight at 4°C with primary antibodies, such as anti-MMP13 (diluted 1:200, Abcam Cat# ab39012, RRID: AB_776416), anti-COL II (diluted 1:1000, Abcam Cat# ab34712, RRID: AB_731688), anti-Aggrecan (diluted 1:200, Abcam Cat# ab36861, RRID: AB_722655) or anti-ADAMTS 5 (diluted 1:500, Abcam Cat# ab41037, RRID: AB_2222327). Then, the cells were incubated with Alexa Flour 488-labelled goat anti-mouse IgG (Invitrogen 1:200; Invitrogen, OR, USA) for 1 h at room temperature. Cells were analysed using an Olympus DP72 fluorescence microscope (Olympus, Tokyo, Japan). To detect NP cell apoptosis, TUNEL staining was performed using a kit based on the manufacturer’s instructions (Promega, Fitchburg, WI, USA).
Alexa flour 488 labelled goat anti mouse igg
Manufactured by Thermo Fisher Scientific
Sourced in United States
Alexa Fluor 488-labelled goat anti-mouse IgG is a secondary antibody reagent used for the detection and visualization of mouse primary antibodies in various immunoassay and imaging applications. It is conjugated with the Alexa Fluor 488 fluorescent dye, which provides bright, photostable green fluorescence.
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2 protocols using alexa flour 488 labelled goat anti mouse igg
1
Immunofluorescence Analysis of NP Cells
2
Immunofluorescence Analysis of NP Cells
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Cultured human NP cells were rinsed with PBS three times, fixed with 4% formaldehyde for 15 min, and permeabilized with 0.3% Triton-X100 (Sigma, CO, USA) for 10 min. Subsequently, the cells were blocked with 6% BSA for 1 h and then were incubated overnight at 4°C with primary antibodies, such as anti-MMP13 (diluted 1:200, Abcam Cat# ab39012, RRID: AB_776416), anti-COL II (diluted 1:1000, Abcam Cat# ab34712, RRID: AB_731688), anti-Aggrecan (diluted 1:200, Abcam Cat# ab36861, RRID: AB_722655) or anti-ADAMTS-5 (diluted 1:500, Abcam Cat# ab41037, RRID: AB_2222327). Then, the cells were incubated with Alexa Flour 488-labelled goat anti-mouse IgG (Invitrogen 1:200; Invitrogen, OR, USA) for 1 h at room temperature. Cells were analysed using an Olympus DP72 fluorescence microscope (Olympus, Tokyo, Japan). To detect NP cell apoptosis, TUNEL staining was performed using a kit based on the manufacturer's instructions (Promega, Fitchburg, WI, USA).
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