Document Detail

An immunofluorescence method to analyze the proliferation status of individual nephron segments in the Xenopus pronephric kidney.
MedLine Citation:
PMID:  22639256     Owner:  NLM     Status:  MEDLINE    
Organ development requires the coordination of proliferation and differentiation of various cell types. This is particularly challenging in the kidney, where up to 26 different cell types with highly specialized functions are present. Moreover, even though the nephron initially develops from a common progenitor pool, the individual nephron segments are ultimately quite different in respect to cell numbers. This suggests that some cells in the nephron have a higher proliferative index (i.e., cell cycle length) than others. Here, we describe two different immunofluorescence-based approaches to accurately quantify such growth rates in the pronephric kidney of Xenopus laevis. Rapidly dividing cells were identified with the mitosis marker phospho-Histone H3, while slowly cycling cells were labeled using the thymidine analogue EdU. In addition, individual nephron segments were marked using cell type-specific antibodies. To accurately assess the number of positively stained cells, embryos were then serially sectioned and analyzed by immunofluorescence microscopy. Growth rates were established by counting the mitosis or S-phase events in relation to the overall cells present in the nephron segment of interest. This experimental design is very reproducible and can easily be modified to fit other animal models and organ systems.
Daniel Romaker; Bo Zhang; Oliver Wessely
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Methods in molecular biology (Clifton, N.J.)     Volume:  886     ISSN:  1940-6029     ISO Abbreviation:  Methods Mol. Biol.     Publication Date:  2012  
Date Detail:
Created Date:  2012-05-28     Completed Date:  2012-09-18     Revised Date:  2013-06-24    
Medline Journal Info:
Nlm Unique ID:  9214969     Medline TA:  Methods Mol Biol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  121-32     Citation Subset:  IM    
Department of Cell Biology, Lerner Research Institute/Cleveland Clinic, Cleveland, OH, USA.
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MeSH Terms
Cell Cycle
Cell Proliferation
Fluorescent Antibody Technique / methods*
Kidney / cytology,  embryology*
Nephrons / cytology*
Pronephros / cytology*,  embryology
Xenopus / embryology*
Grant Support

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