Cell kinetic analysis of intact rat colonic crypts by confocal microscopy and immunofluorescence

Gastroenterology
H KonishiN H Terry

Abstract

Precise quantitative and spatial analysis of cell cycle-related biomarkers in colonic crypts is often vital for studies of colon carcinogenesis and cancer prevention. To overcome the limitations of histology, confocal laser microscopy of microdissected whole crypts was used to quantitate S phase and mitotic cells. Microdissected distal colonic crypts were studied in a modified rat starvation refeeding model. S phase cells were labeled in vivo with 5-bromodeoxyuridine. Mitotic cells were labeled with MPM2 (antibody to mitosis-specific epitope) and also assessed for chromatin morphology with propidium iodide. Sequential optical crypt sections, produced by confocal microscopy, were digitally imaged. S phase labeling indices per whole crypt were also compared with those derived by conventional immunohistochemistry. S phase and mitotic cells were clearly discriminated without background staining. The labeled S phase cell number and fraction per whole crypt were significantly decreased with starvation and increased with refeeding. Variability in the labeling index between whole crypts analyzed by confocal microscopy was significantly smaller than between histological crypt sections. Consequently, the intervention contributed to 92.2%...Continue Reading

Citations

Dec 13, 2005·Surgery Today·Shinya OdaYoshihiko Maehara
Mar 11, 2006·Familial Cancer·Jacques BertholonAlain Puisieux
Oct 13, 1998·In Vitro Cellular & Developmental Biology. Animal·B Kaeffer, S Briollais
Jul 10, 1998·The Proceedings of the Nutrition Society·J C Mathers
Feb 28, 2003·In Vitro Cellular & Developmental Biology. Animal·Bertrand KaefferChristine Cherbut
Jan 6, 2006·Chronobiology International·L PardiniJ-P Galmiche

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