Cilia were immunostained with anti-Arl13b antibody (T. Caspary), and cilia size was determined by dividing the number of Arl13b-positive pixels by the number of nuclei per image. Ciliary levels of Gli2 and IFT88 were determined by immunostaining cells with anti-Gli2 (R&D Systems) or anti-IFT88 (ProteinTech Group) antibodies. Analyses of mitotic spindles and kinetochore-microtubule attachments in fixed cells were performed as described26 (link), and real-time confocal microscopy was conducted with NIH-3T3 cells stably expressing GFP-tubulin. Melanosome and peroxisome motility studies were conducted as reported19 (link),20 (link).
Protocol detail
Analyzing Cellular Organelle Dynamics
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Anti antibody
Antibodies
Cells
Cells attachments
Cilia
Confocal microscopy
Kinetochore
Levels
Melanosome
Microtubule
Mitotic spindles
Motility
Nih 3t3 cells
Nuclei
Peroxisome
Tubulin
Corresponding Organization :
Other organizations : Stanford University, Rockefeller University, Northwestern University
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Protocol cited in 8 other protocols
Variable analysis
independent variables
- Anti-Arl13b antibody
- Anti-Gli2 antibody
- Anti-IFT88 antibody
- Cilia size
- Ciliary levels of Gli2
- Ciliary levels of IFT88
- Mitotic spindle
- Kinetochore-microtubule attachments
- Melanosome motility
- Peroxisome motility
- Number of nuclei per image
- NIH-3T3 cells stably expressing GFP-tubulin
- Positive control: Not specified.
- Negative control: Not specified.
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