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Abstract:

The flash photolysis of "caged" compounds is a powerful experimental technique for producing rapid changes in concentrations of bioactive signaling molecules. These caged compounds are inactive and become active when illuminated with ultraviolet light. This paper describes an inexpensive adaptation of an Olympus confocal microscope that uses as source of ultraviolet light the mercury lamp that comes with the microscope for conventional fluorescence microscopy. The ultraviolet illumination from the lamp (350 - 400 nm) enters through an optical fiber that is coupled to a nonconventional port of the microscope. The modification allows to perform the photolysis of caged compounds over wide areas (~200 μm) and obtain confocal fluorescence images simultaneously. By controlling the ultraviolet illumination exposure time and intensity it is possible to regulate the amount of photolyzed compounds. In the paper we characterize the properties of the system and show its capabilities with experiments done in aqueous solution and in Xenopus Laevis oocytes. The latter demonstrate its applicability for the study of Inositol 1,4,5-trisphosphate-mediated intracellular calcium signals. © 2011 Society of Photo-Optical Instrumentation Engineers (SPIE).

Registro:

Documento: Artículo
Título:Custom-made modification of a commercial confocal microscope to photolyze caged compounds using the conventional illumination module and its application to the observation of Inositol 1,4,5-trisphosphate-mediated calcium signals
Autor:Sigaut, L.; Barella, M.; Espada, R.; Ponce, M.L.; Dawson, S.P.
Filiación:Ciudad Universitaria, Departamento de Física, FCEN-UBA, and IFIBA, CONICET, Pabell ón I, 1428 Buenos Aires, Argentina
Palabras clave:Cage compound; Calcium signal; Confocal microscopy; Inositol 1,4,5-trisphosphate receptor; Photolysis; UV illumination; Cage compounds; Confocal fluorescence; Confocal microscopes; Experimental techniques; Exposure-time; Flash photolysis; Inositol 1,4,5-trisphosphate receptor; Intracellular calcium; Olympus; Rapid changes; Signaling molecules; Ultra-violet light; Ultraviolet illumination; UV illuminations; Wide area; Xenopus laevis oocytes; Alcohols; Calcium; Confocal microscopy; Enzyme activity; Fluorescence; Fluorescence microscopy; Mercury (metal); Optical fibers; Photolysis; Sugars; Ultraviolet radiation; Microscopes; Xenopus laevis; 2 nitrophenyl EGTA; 2-nitrophenyl-EGTA; calcium; drug derivative; egtazic acid; inositol 1,4,5 trisphosphate; animal; article; calcium signaling; chemistry; confocal microscopy; equipment design; instrumentation; metabolism; methodology; oocyte; photolysis; physiology; statistical model; ultraviolet radiation; Xenopus laevis; Animals; Calcium; Calcium Signaling; Egtazic Acid; Equipment Design; Inositol 1,4,5-Trisphosphate; Linear Models; Microscopy, Confocal; Oocytes; Photolysis; Ultraviolet Rays; Xenopus laevis
Año:2011
Volumen:16
Número:6
DOI: http://dx.doi.org/10.1117/1.3592497
Título revista:Journal of Biomedical Optics
Título revista abreviado:J Biomed Opt
ISSN:10833668
CODEN:JBOPF
CAS:calcium, 14092-94-5, 7440-70-2; egtazic acid, 67-42-5; inositol 1,4,5 trisphosphate, 85166-31-0, 88269-39-0; 2-nitrophenyl-EGTA, 238073-40-0; Calcium, 7440-70-2; Egtazic Acid, 67-42-5; Inositol 1,4,5-Trisphosphate, 85166-31-0
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10833668_v16_n6_p_Sigaut

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Citas:

---------- APA ----------
Sigaut, L., Barella, M., Espada, R., Ponce, M.L. & Dawson, S.P. (2011) . Custom-made modification of a commercial confocal microscope to photolyze caged compounds using the conventional illumination module and its application to the observation of Inositol 1,4,5-trisphosphate-mediated calcium signals. Journal of Biomedical Optics, 16(6).
http://dx.doi.org/10.1117/1.3592497
---------- CHICAGO ----------
Sigaut, L., Barella, M., Espada, R., Ponce, M.L., Dawson, S.P. "Custom-made modification of a commercial confocal microscope to photolyze caged compounds using the conventional illumination module and its application to the observation of Inositol 1,4,5-trisphosphate-mediated calcium signals" . Journal of Biomedical Optics 16, no. 6 (2011).
http://dx.doi.org/10.1117/1.3592497
---------- MLA ----------
Sigaut, L., Barella, M., Espada, R., Ponce, M.L., Dawson, S.P. "Custom-made modification of a commercial confocal microscope to photolyze caged compounds using the conventional illumination module and its application to the observation of Inositol 1,4,5-trisphosphate-mediated calcium signals" . Journal of Biomedical Optics, vol. 16, no. 6, 2011.
http://dx.doi.org/10.1117/1.3592497
---------- VANCOUVER ----------
Sigaut, L., Barella, M., Espada, R., Ponce, M.L., Dawson, S.P. Custom-made modification of a commercial confocal microscope to photolyze caged compounds using the conventional illumination module and its application to the observation of Inositol 1,4,5-trisphosphate-mediated calcium signals. J Biomed Opt. 2011;16(6).
http://dx.doi.org/10.1117/1.3592497