Measuring the secretion of heat shock proteins from cells.
Heat shock proteins have been shown to be secreted from a number of cell types. Necrotic cells release heat shock proteins in a passive manner, whereas we, and others, have shown that viable cells secrete Hsp70 and Hsp60 through an active mechanism involving lysosomal vesicles and lipid rafts. This release of Hsp70 and Hsp60 is regulated, for example by being increased by elevated temperature. This article outlines procedures, using Hsp70 as the example, to: ensure the status of cells (viable, apoptotic or necrotic); identify the heat shock protein secreted; and quantify the secreted protein. Hsp70 has previously been quantified by ELISA, but newer methods are now being adopted, such as BIAcore and bead-based assays for use by FACS. These methods have the advantages of being more sensitive and requiring less sample than ELISA. The BIAcore has the potential to analyse Hsp70 ligands and provide affinity constants. The FACS bead assay system can be used to run multiplex assays.
Ireland
H Elyse
HE
Chester Centre for Stress Research, Department of Biological Sciences, University of Chester, Parkgate Road, Chester CH1 4BJ, UK.
Leoni
Francesca
F
Altaie
Ola
O
Birch
Catherine S
CS
Coleman
Robert C
RC
Hunter-Lavin
Claire
C
Williams
John H H
JH
D000818:Animals; D002462:Cell Membrane / Q000473:pathology; D002468:Cell Physiological Phenomena; D002470:Cell Survival; D005434:Flow Cytometry; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008214:Lymphocytes / Q000166:cytology / Q000502:physiology; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D009336:Necrosis; D013816:Thermodynamics
D006360:Heat-Shock Proteins