Immunofluorescent staining was performed as described previously [49 (link)]. Cells were fixed in 4% paraformaldehyde, permeabilized with ice-cold 100% methanol, and immunostained with LC3B and LAMP2 (ab25631; Abcam, Cambridge, MA, USA) antibodies followed by incubation with CF488A-conjugated goat-anti-mouse IgG (#20018) and CF568-conjugated goat-anti-rabbit IgG (#20103), highly cross-absorbed (Biotium, Hayward, CA, USA). Nuclei were revealed by Hoechst 33342 staining. Fluorescence images were collected under a Leica DMIRB fluorescent microscope (Leica Microsystems, Wetzlar, Germany) equipped with a Spinning Disk Confocal Microscopy system (UltraView cooled CCD; Perkin Elmer, Waltham, MA, USA).
Dmirb fluorescent microscope
Manufactured by Leica
Sourced in Germany
The Leica DMIRB is a fluorescent microscope designed for advanced imaging applications. It features a high-performance illumination system and precision optics to capture detailed fluorescent images of biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Ultraview cooled ccd, by PerkinElmer (4 mentions)
Cf568 conjugated goat anti rabbit igg, by Biotium (3 mentions)
Cf488 conjugated goat anti mouse igg, by Biotium (2 mentions)
Cf488a conjugated goat anti mouse igg, by Biotium (1 mentions)
Mouse anti β actin, by Cell Signaling Technology (1 mentions)
Rabbit anti vasp, by Cell Signaling Technology (1 mentions)
Cf488 conjugated goat anti rabbit igg, by Biotium (1 mentions)
4 protocols using dmirb fluorescent microscope
1
Immunofluorescent Imaging of Autophagy Markers
2
Immunofluorescent Visualization of Cytoskeletal Proteins
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After incubation with CuB (0.1 μM), cells were fixed in 4% paraformaldehyde prepared in phosphate-buffered saline (PBS), permeabilized with ice-cold 100% methanol, and immunostained with mouse anti-β-actin (1∶500; Cell Signaling Technology) or rabbit anti-VASP (1∶300; Cell Signaling Technology), followed by CF488-conjugated goat-anti-mouse IgG (1∶700) or CF568-conjugated goat-anti-rabbit IgG (1∶700), highly cross-absorbed (Biotium, Hayward, CA, USA). Nuclei were revealed by Hoechst33342 (5 μg/ml) staining. Fluorescence images were observed and collected under a Leica DMIRB fluorescent microscope (Leica Microsystems, Wetzlar, Germany) armed with a Spinning Disk Confocal Microscopy system (UltraView cooled CCD; Perkin Elmer, Waltham, MA, USA).
3
Immunofluorescence Analysis of Cytoskeletal and Transcription Factor
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Immunofluorescence analysis was performed as previously described [16] (link). Cells were fixed in 4% paraformaldehyde, permeabilized with ice-cold 100% methanol, and immunostained with mouse anti-β-actin and rabbit anti-p65 antibodies followed by CF488-conjugated goat-anti-mouse IgG and CF568-conjugated goat-anti-rabbit IgG, highly cross-absorbed (Biotium, Hayward, CA). Nuclei were revealed by Hoechst 33342 staining. Fluorescence images were collected under a Leica DMIRB fluorescent microscope (Leica Microsystems, Wetzlar, Germany) armed with a Spinning Disk Confocal Microscopy system (UltraView cooled CCD; Perkin Elmer, Waltham, MA, USA).
4
Immunofluorescence Staining of U251 Cells
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U251cells were fixed with 4% paraformaldehyde and permeabilized with 100% methanol. 10% normal goat serum was used to block the nonspecific binding with primary antibodies. Then cells were incubated with appropriate primary antibodies at 4o C overnight. After PBS wash, cells were incubated with CF568-conjugated goat-anti-mouse IgG or CF488-conjugated goat-anti-rabbit IgG (Biotium, Hayward, CA, USA) at room temperature for 1 hr. Nuclei were revealed by Hoechst33342 staining. Fluorescence images were collected under a Leica DMIRB fluorescent microscope (Leica Microsystems, Wetzlar, Germany) armed with a spinning disk confocal microscopy system (UltraView cooled CCD; Perkin Elmer, Waltham, MA, USA).
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