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A specific fluorogenic reaction for tryptophan residues using isatin in organic solvents
C I Trigoso1, N Ibañez, J C Stockert
1Department of Biology, Faculty of Sciences, Autonomous University of Madrid, Spain.
Summary
Isatin is a novel fluorogenic reagent that specifically detects tryptophan residues in cells. This method enables bright green fluorescence in eosinophil leucocyte granules, aiding cytochemistry research.
Area of Science:
- Biochemistry
- Cell Biology
- Analytical Chemistry
Background:
- Fluorogenic reactions are crucial for specific chemical group detection in cytochemistry.
- Isatin, a nonfluorescent compound, has potential applications in biological staining.
Purpose of the Study:
- To explore the use of isatin as a fluorogenic reagent for identifying specific chemical groups in fixed biological samples.
- To investigate the specificity of isatin's reaction with cellular components.
Main Methods:
- Methanol-fixed blood smears were treated with 0.1% isatin in ethanol or acetone.
- Samples were observed under violet-blue exciting light.
- Tryptophan residues were blocked using performic acid oxidation to assess reaction specificity.
Main Results:
- Isatin treatment induced bright green fluorescence in eosinophil leucocyte granules.
- The fluorescence was abolished after blocking tryptophan residues.
- In vitro spectrofluorometric studies confirmed that isatin forms a unique fluorescent product specifically with tryptophan.
Conclusions:
- Isatin serves as a specific fluorogenic reagent for detecting tryptophan residues.
- This reaction provides a valuable tool for cytochemical analysis, particularly for identifying eosinophil granules.