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. 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.
Department of Pharmacology and Toxicology, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA. kasemn@mcw.edu
Kasem
K
Nithipatikom
Michael J
MJ
McCoy
Sharon R
SR
Hawi
Kazuo
K
Nakamoto
Fran
F
Adar
William B
WB
Campbell
D000804:Angiotensin II / Q000032:analysis; D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000032:analysis; D048588:Benzoxazines; D001710:Biotin / Q000737:chemistry; D002417:Cattle; D002478:Cells, Cultured; D002951:Citrates / Q000737:chemistry; D003102:Colloids / Q000737:chemistry; D004396:Coloring Agents / Q000737:chemistry*; D003331:Coronary Vessels / Q000166:cytology / Q000201:enzymology; D051545:Cyclooxygenase 1; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology; D005455:Fluorescent Antibody Technique / Q000379:methods; D007527:Isoenzymes / Q000032:analysis; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods; D010078:Oxazines / Q000737:chemistry; D011451:Prostaglandin-Endoperoxide Synthases / Q000032:analysis; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D011945:Receptors, Angiotensin / Q000378:metabolism; D012680:Sensitivity and Specificity; D018030:Silver Compounds / Q000737:chemistry; D013059:Spectrum Analysis, Raman / Q000379:methods*; D015384:Zona Glomerulosa; D004124:p-Dimethylaminoazobenzene / Q000737:chemistry
D048588:Benzoxazines; D002951:Citrates; D003102:Colloids; D004396:Coloring Agents; D007527:Isoenzymes; D010078:Oxazines; D011506:Proteins; D011945:Receptors, Angiotensin; D018030:Silver Compounds; D000804:Angiotensin II; C028911:cresyl violet; D001710:Biotin; D004124:p-Dimethylaminoazobenzene; D001092:Arachidonate 12-Lipoxygenase; D051545:Cyclooxygenase 1; D011451:Prostaglandin-Endoperoxide Synthases