Document Detail


A novel approach for monitoring extracellular acidification rates: based on bead injection spectrophotometry and the lab-on-valve system.
MedLine Citation:
PMID:  14978521     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Monitoring extracellular acidification rates (ECARs) is important for the study of cellular activities, since it allows for the evaluation of factors that alter metabolic function, such as stimulants, inhibitors, toxins as well as receptor and non-receptor mediated events. While the light addressable potentiometric sensor (Cytosensor Microphysiometer) has been the principal tool for ECARs measurement in the past, this work introduces a novel method that exploits an immobilized pH indicator on the surface of microcarrier beads (Sephadex) and is probed with a fiber optic coupled spectrophotometer. Likewise, live cells under investigation were also immobilized on microcarrier beads (Cytopore). These beads are metered, transported and monitored within a microfluidic system, termed as the Lab-on-Valve (LOV). Use of carrier beads in conjunction with Bead Injection Spectrophotometry and a Lab-on-Valve module (BIS-LOV), makes ECAR measurements reliable and automated. The feasibility of the BIS-LOV approach is demonstrated measuring ECARs of the mouse hepatocyte cell line, TABX.2S, grown on Cytopore beads packed within the central channel of the LOV system. These immobilized cells were perfused in a phosphate buffer carrier solution (capacity: 1 mmol L(-1), pH 7.4). Protons extruded from 10(5) to 10(6) cells were accumulated during a stopped flow period of 220 s followed by a pH measurement, detected by changes in absorbance of the pH indicator bonded to the microcarrier beads. Addition of metabolic inhibitors (sodium azide, oxamic acid) to the carrier buffer solution can induced an increase or decrease of the basal proton extrusion rate in a very reproducible manner. Comparison of the BIS-LOV technique to the Cytosensor microphysiometer and literature confirms the validity of this novel approach, highlighting its advantages and suggesting future improvements that will make the BIS-LOV a practical tool for routine ECARs measurement.
Authors:
Holger A Erxleben; Michael K Manion; David M Hockenbery; Louis Scampavia; Jaromir Ruzicka
Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.     Date:  2004-02-02
Journal Detail:
Title:  The Analyst     Volume:  129     ISSN:  0003-2654     ISO Abbreviation:  Analyst     Publication Date:  2004 Mar 
Date Detail:
Created Date:  2004-02-23     Completed Date:  2004-05-21     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0372652     Medline TA:  Analyst     Country:  England    
Other Details:
Languages:  eng     Pagination:  205-12     Citation Subset:  IM    
Affiliation:
Department of Chemistry, Box 351700, University of Washington, Seattle, WA 98195-1700, USA.
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MeSH Terms
Descriptor/Qualifier:
Animals
Cell Line
Extracellular Space / metabolism*
Flow Injection Analysis
Hepatocytes / metabolism*
Hydrogen-Ion Concentration
Mice
Microchemistry
Spectrophotometry, Ultraviolet
Grant Support
ID/Acronym/Agency:
5R01 EB 000782-12/EB/NIBIB NIH HHS; 5T32 CA 08416/CA/NCI NIH HHS; U01 CA 91310/CA/NCI NIH HHS

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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