HeLa cells were seeded into Lab-Tek II 8-well chamber slides (CC2 Glass slide, Nunc; Rochester, NY). After 16 h, 200ng of HA-UB and NS1 plasmids were transfected with Lipofectamine 2000 for 30 h, followed by treatment with MG-132 (20 mM) (or DMSO as control) for 6 h. The cells were washed with DPBS 1X, fixed with 4% paraformaldehyde, permeabilized with 0.5% NP-40 (v/v) in DPBS 1X, and blocked with 0.3% BSA, 0.2% fish gelatin in DPBS 1X for 1 h (blocking solution). Staining was performed with anti-HA and anti-NS1 antibodies (1:200) (prepared in blocking solution) overnight at 4°C. The next day, cells were washed three times with DPBS 1X and incubated with the secondary antibodies donkey anti-rabbit Alexa-fluor 488 and anti-mouse 555 (Invitrogen) diluted in blocking buffer along with DAPI (1:2000). GFP and RFP fluorescence-positive cells were imaged by microscopy under a 60X objective lens on a Cytation 5 Imaging reader (Biotek Instruments).
Lab tek 2 8 well chamber slides cc2 glass slide
Manufactured by Thermo Fisher Scientific
Lab-Tek II 8-well chamber slides (CC2 Glass slide) are a laboratory product designed for cell culture and microscopic observation. The slides provide a contained environment for culturing cells and enable their direct visualization under a microscope.
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2 protocols using lab tek 2 8 well chamber slides cc2 glass slide
1
Imaging Protein Interactions in HeLa Cells
2
Immunofluorescence Analysis of TRIM21 and HBV Pol
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Huh7 cells were seeded into Lab-Tek II 8-well chamber slides (CC2 Glass slide, Nunc; Rochester, NY). After 18 h, 500 ng of HA-TRIM21 and FLAG-HBV DNA Pol plasmids were transfected with Lipofectamine 2000 for 36 h. The cells were washed with 1×PBS, fixed with 4% paraformaldehyde in PBS for 30 min at 37 °C, and permeabilized with 0.5% Triton X-100 (v/v), followed by blocking in PBS containing 10% Donkey serum (Solarbio, Shanghai, China) at room temperature for 2 h. Cells were incubated with anti-HA (1:100) and anti-FLAG antibodies (1:500) diluted in blocking buffer overnight at 4°C. The next day, cells were washed three times with 1×PBS and incubated with the secondary antibodies donkey anti-rabbit and anti-mouse (Invitrogen) diluted in blocking buffer in the dark. Then, cells were mounted with DAPI Fluoromount-G (Southern Biotech, Birmingham, Alabama, USA), and images were captured with an LSM 700 confocal microscope (Zeiss, Thornwood, NY, USA).
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