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A Label-free Technique for the Spatio-temporal Imaging of Single Cell Secretions
Published on: November 24, 2015
[Study of solid-phase time-resolved fluorescence label immunoassay]
Li-hua Pan1, Si-hong Zhou, Wen-wei Sun
1National Analytical Research Center of Electrochemistry and Spectroscopy, Key Laboratory of Rare Earth Chemistry and Physics, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China.
Insights
This study optimizes labeling of Hepatitis B surface antibody IgG (HBsAbIgG) using a novel fluorescence probe (BCPDA) for time-resolved fluorimmunoassays. The method ensures high bioactivity and stable, intense fluorescence for improved diagnostics.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Immunodiagnostics
Context:
- Solid-phase time-resolved fluorimmunoassays (TRFIA) are crucial for sensitive detection.
- Hepatitis B surface antibody IgG (HBsAbIgG) detection is vital for diagnosing Hepatitis B virus infections.
- Developing efficient labeling methods is key to enhancing immunoassay performance.
Purpose:
- To determine optimal conditions for labeling HBsAbIgG with 4,7-bis-chorosulfophenyl-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA).
- To characterize the properties of the resulting protein-fluorophore complex for TRFIA applications.
Summary:
- BCPDA reacts with HBsAbIgG under mild conditions, preserving over 80% bioactivity.
- Optimal BCPDA to HBsAbIgG molar ratio is 45-70, with high recovery (>80%).
- The stable Protein-BCPDA-Eu3+ complex exhibits high fluorescence intensity, long lifetimes, a large Stokes shift (270 nm), and a narrow emission band (611.2 nm).
Impact:
- Provides a foundation for developing non-isotopic immunoassay techniques.
- Facilitates advancements in clinical medical diagnosis for Hepatitis B.
- Offers a highly sensitive and stable fluorescent labeling strategy for biomolecules.
Abstract:
This paper describes optimal conditions for HBsAbIgG labeling with a new fluorescence probe, 4,7-bis-chorosulfophenyl-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA) for the solid phase time-resolved fluorimmunoassay (TRFIA). The result of experiment under states clearly that BCPDA may react with protein under relative mild condition. The relative bioactivity of reacted protein was more than 80%. The labeling molar ratio of BCPDA for HBsAbIgG was 45-70. The recovery was higher than 80%. Protein-BCPDA-Eu3+ complex is stable. It can emit very high fluorescence intensity with very long fluorescence life times. The fluorescence of Protein-BCPDA-Eu3+ complex has a very large stokes shift (270 nm). The emission band at 611.2 nm is very narrow. The research provides the base for developing non-isotopic immunoassay technique and clinical medical diagnosis.
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