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1. Introduction
2. ROS/RNS-induced radical cascade
3. Overview
4. Assays for ROS and RNS
5. Similarity between lucigenin and paraquat radicals
6. Lucigenin: chemiluminescent probe for superoxide
7. Spin trapping technique
8. Properties of a "good" spin trap
9. Most commonly used cyclic nitrone spin trap
10. DMPO spin-trapping
11. Newly-synthesized phosphorylated spin trap
12. DEPMPO spin-trapping
13. Newly-synthesized carboxylated trap
14. EMPO spin-trapping
15. Relative half-lives of superoxide adducts
16. New solid cyclic nitrone trap
17. BMPO spin-trapping
18. Spin-trapping of superoxide with BMPO trap
19. Spin-trapping of hydroxyl radical with BMPO trap
20. Mitochondria targeted spin traps
21. Oxidative injury in vascular diseases
22. Activation of endothelial nitric oxide synthase
23. Generation of superoxide by eNOS
24. BH4 inhibits superoxide generation from eNOS
25. Probes for oxidation states
26. What can we study using fluorescent probes?
27. Oxidation of DCFH by hydrogen-peroxide and iron
28. Detecting oxidizing species (non-specific)
29. Measuring intracellular oxidative stress by DCF
30. Intracellular oxidation of DCFH to DCF
31. DCFH oxidation depends on intracellular GSH
32. DCFH oxidation depends on iron signaling
33. Reaction between hydroethidine and superoxide
34. Fluorescence spectra of HE oxidation product
35. Does superoxide react with HE to form ethidium?
36. Superoxide generating systems
37. HE/X/XO-DNA and EB-DNA fluorescence spectra
38. HE/KO2-DNA and EB-DNA fluorescence spectra
39. HE-BH4-free eNOS-DNA and EB-DNA spectra
40. HE/superoxide, EB-DNA reaction product spectra
41. HE-X/XO product: liquid chromatography
42. Mass spectrometry
43. HE and superoxide reaction - summary
44. HE and superoxide reaction - conclusion
45. Determining the HE/superoxide product structure
46. Structure of the HE/superoxide reaction product
47. Menadione-induced HE fluorescence in BAEC
48. HPLC identification of 2-hydroxyethidium
49. HPLC calibration using authentic compounds
50. Intracellular concentration of 2-OH-E+ and E+
51. The effect of SOD on product formation
52. Comparison between HPLC and EPR analyses
53. HPLC analysis of superoxide production in cells
54. Optimal conditions for detection by fluorescence
55. 2-hydroxyethidium formation - conclusions
56. Independent synthesis of 2-OH-E+
57. Chemical structures of HE, Fs and NDS
58. HPLC analysis of the HE/Fremy's salt reaction
59. HPLC/mass spectrum of the reaction product
60. Stoichiometry of HE and Fremy's salt reaction
61. Mechanism of oxidation of HE to 2-OH-E+
62. HPLC/EC detection of HE, E+, and 2-OH-E+
63. Detection of cellular nitric-oxide by fluorescence
64. DAF-2 fluorescence and DAF-2T formation
65. HPLC of DAF-2 and DAF-2T
66. Relevant publications
67. Acknowledgement

Topics Covered

• Reactive oxygen and nitrogen species play a critical role in physiological and pathological signaling processes
• Because of the rapid reaction between the radicals (for example, nitric oxide and superoxide), it is very difficult to detect and quantitate the formation of these species under pathophysiological conditions
• Complex interrelationship between the reactive oxygen and nitrogen species
• Importance of understanding the chemistry of the fluorophores and spin traps that are often employed in the detection of these species

Links

Series:

• Free Radicals and the Oxygen Paradox

Categories:

• Biochemistry

Talk Citation

Kalyanaraman, B. (2007, November 1). Chemistry and biochemistry of molecular probes used in the detection of reactive oxygen and nitrogen species [Video file]. In The Biomedical & Life Sciences Collection, Henry Stewart Talks. Retrieved April 20, 2024, from https://hstalks.com/bs/376/.
Export Citation (RIS)

Publication History

• Published on November 1, 2007
• Archived on May 31, 2018

Financial Disclosures

• Prof. Balaraman Kalyanaraman has not informed HSTalks of any commercial/financial relationship that it is appropriate to disclose.