Rosa26-Tomato females were crossed with Hoxb7Cre;EF1aCrb3-GFP/+ males and kidneys were dissected out at E13.5 (Madisen et al., 2010 (link)). Kidneys were plated on a thin Matrigel layer (1:4 in medium; BD Biosciences, Franklin Lakes, NJ) in Lab-Tek Chambered #1.0 Borosilicate Coverglass System dishes (Nalge Nunc International, Rochester, NY). Additional medium was added to surround half of the kidney, allowing growth at the air/media interface. Four kidneys were cultured and two of them were imaged at 20× magnification. Pictures were taken every 20 min for 20 h using a Nikon A1 laser scanning confocal microscope with a Tokai Hit stage Top incubator (Tokai Hit Co., Shizuoka, Japan).
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Kidney Development Imaging Protocol
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Corresponding Organization : The University of Texas Southwestern Medical Center
Other organizations : Washington University in St. Louis
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Variable analysis
independent variables
- Crossing Rosa26-Tomato females with Hoxb7Cre;EF1aCrb3-GFP/+ males
- Kidney growth and development observed over 20 hours
- Matrigel layer (1:4 in medium) used for plating the kidneys
- Lab-Tek Chambered #1.0 Borosilicate Coverglass System dishes used for culturing the kidneys
- Allowing growth at the air/media interface
- Imaging using a Nikon A1 laser scanning confocal microscope with a Tokai Hit stage Top incubator
- No positive or negative controls were explicitly mentioned in the input protocol.
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