Characterization and application of Raman labels for confocal Raman microspectroscopic detection of cellular proteins in single cells

A method using confocal Raman microspectroscopy for the detection of cellular proteins in single intact cells was developed. Two approaches were used to improve the detection of these cellular components. First, compounds with high Raman scattering were investigated for potential use as Raman labels...

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Bibliographic Details
Published in:	Analytical biochemistry 2003-11, Vol.322 (2), p.198-207
Main Authors:	Nithipatikom, Kasem, McCoy, Michael J, Hawi, Sharon R, Nakamoto, Kazuo, Adar, Fran, Campbell, William B
Format:	Article
Language:	eng
Subjects:	Angiotensin II Angiotensin II - analysis Animals Arachidonate 12-Lipoxygenase - analysis Benzoxazines Biotin - chemistry Cattle Cells, Cultured Citrates - chemistry Colloids - chemistry Coloring Agents - chemistry Coronary Vessels - cytology Coronary Vessels - enzymology Cyclooxygenase Cyclooxygenase 1 Endothelium, Vascular - cytology Endothelium, Vascular - enzymology Fluorescent Antibody Technique - methods Isoenzymes - analysis Lipoxygenase Microscopy, Confocal - instrumentation Microscopy, Confocal - methods Oxazines - chemistry p-Dimethylaminoazobenzene - chemistry Prostaglandin-Endoperoxide Synthases - analysis Proteins - analysis Proteins - chemistry Raman labels Receptors Receptors, Angiotensin - metabolism Sensitivity and Specificity Silver Compounds - chemistry Spectrum Analysis, Raman - methods Surface-enhanced Raman scattering Zona Glomerulosa
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Summary:	A method using confocal Raman microspectroscopy for the detection of cellular proteins in single intact cells was developed. Two approaches were used to improve the detection of these cellular components. First, compounds with high Raman scattering were investigated for potential use as Raman labels. Raman labels were conjugated to either biomolecules or biotin and used as markers in the detection of cellular enzymes and receptors. Second, silver colloids were used to increase the surface-enhanced Raman scatter (SERS) of these Raman labels. Cresyl violet and dimethylaminoazobenzene are Raman labels that provide very sensitive SERS detection by a confocal Raman microscope with a HeNe laser at wavelength of 632.8 nm. The detection of 12-lipoxygenase and cyclooxygenase-1 in single bovine coronary artery endothelial cells and the binding of angiotensin II to its receptors in zona glomerulosa cells was demonstrated.
ISSN:	0003-2697 1096-0309