Document Detail


Comparison of lymphocyte isolation methods for endoscopic biopsy specimens from the colonic mucosa.
MedLine Citation:
PMID:  23279944     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
An ideal method of immune cell isolation should provide maximum cell yield without disturbing functional properties. Intestinal endoscopic biopsies, in contrast to surgical samples, allow the study of all disease stages but have the drawback of a minimum amount of tissue available, making protocol optimization mandatory. We compared for the first time two methods of separation of colonic epithelium and five methods of lamina propria cell isolation for colonic biopsy specimens (mechanical, enzymatic and organ culture protocols). Lymphocyte number, viability and phenotype (CD45+, CD103+, CD3+, CD4+, CD8+, CD19+, CD16-56) were analyzed by flow cytometry. Neither of the two epithelial detachment protocols achieved proper epithelial separation, though the high intensity ion chelation method was more accurate. Maximum cell yield of lamina propria lymphocytes without phenotypic modification was obtained with overnight smooth enzymatic digestion. High dose collagenase incubation caused a marked decrease in CD4+ lymphocytes of the lamina propria as compared to low enzymatic method (p=0.004). Mechanical and biopsy culture are not advisable methods because of the low cell yield, and phenotypic alterations and high contamination rate, respectively.
Authors:
Anna Carrasco; Josep Mañe; Rebeca Santaolalla; Elisabet Pedrosa; Josefina Mallolas; Violeta Lorén; Marco Fernandez; Fernando Fernández-Bañares; Mercè Rosinach; Carme Loras; Montserrat Forné; Xavier Andújar; Judith Vidal; Josep Maria Viver; Maria Esteve
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-12-29
Journal Detail:
Title:  Journal of immunological methods     Volume:  -     ISSN:  1872-7905     ISO Abbreviation:  J. Immunol. Methods     Publication Date:  2012 Dec 
Date Detail:
Created Date:  2013-1-2     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  1305440     Medline TA:  J Immunol Methods     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
Copyright © 2012. Published by Elsevier B.V.
Affiliation:
Department of Gastroenterology, Hospital Universitari Mútua de Terrassa, University of Barcelona, Research Foundation Mútua de Terrassa, Terrassa, Barcelona, Catalonia, Spain; Department of Pathology , Hospital Universitari Mútua de Terrassa - University of Barcelona - Research Foundation Mútua de Terrassa, Terrassa, Barcelona, Catalonia, Spain; CIBERehd, Centro de Investigación Biomédica en Red en el Área temática de Enfermedades Hepáticas y Digestivas, Spain.
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