Fluorescent immunolabeling of embryonic kidney samples.
This chapter provides a basic protocol to perform fluorescent immunolabeling on embryonic kidney samples. The procedure can be summarized in five steps: permeabilization, primary antibody incubation, washes, secondary antibody incubation, and final washes. This protocol can be used on samples of different origins, from thin sections to whole mounts, just by adjusting incubation times, temperatures, and buffer composition. Despite its simplicity, we have successfully and consistently used this protocol to detect the "usual suspects" in kidney development: CalbindinD-28k, E-cadherin, Pax2, podocalyxin, α-SMA, and Phospho-Histone H3 among others. This protocol also provides a starting point when trying to optimize labeling with a new antibody.
Department of Genetics and Development, Columbia University, New York, NY, USA. cc2636@columbia.edu
Cristina
C
Cebrián
D000818:Animals; D000906:Antibodies / Q000032:analysis; D005455:Fluorescent Antibody Technique / Q000379:methods*; D007668:Kidney / Q000196:embryology*; D008856:Microscopy, Fluorescence / Q000379:methods; D010539:Permeability
D000906:Antibodies