Development of a fluorescent capillary biosensor based on self-assembled AuNP/LDH for micro-volume intracellular...

Area of Science:

• Analytical Chemistry
• Biomedical Engineering
• Biochemistry

Background:

• Intracellular pyruvate (PA) is a key metabolite in cellular energy production.
• Accurate quantification of micro-volume intracellular PA is crucial for understanding metabolic processes.
• Existing methods for PA detection can be limited by sensitivity, sample volume, or reusability.

Purpose of the Study:

• To develop a novel fluorescent capillary biosensor for sensitive and quantitative detection of intracellular pyruvate (PA).
• To optimize the biosensor for micro-volume sample analysis and evaluate its performance characteristics.
• To demonstrate the biosensor's utility for measuring intracellular PA in human erythrocytes.

Main Methods:

• Modification of an amination capillary with gold nanoparticles (AuNPs) and lactate dehydrogenase (LDH) via self-assembly.
• Quantification of PA based on the fluorescence change of NADH in a reaction system.
• Investigation and optimization of various parameters including pH, temperature, reagent concentrations, and reaction times.

Main Results:

• The developed biosensor exhibited a linear response range for PA from 2.5-120 μmol L⁻¹.
• Determination and detection limits were as low as 2.5 and 0.75 μmol L⁻¹, respectively.
• The biosensor demonstrated high reusability (>41 cycles with <1.5% RSD) and achieved 95-106% standard addition recovery.

Conclusions:

• The AuNP-modified fluorescent capillary biosensor offers a sensitive, stable, and reusable platform for intracellular PA quantification.
• The use of AuNPs enhanced LDH affinity, stability, and overall biosensor performance.
• The biosensor successfully measured intracellular PA in human erythrocytes, highlighting its potential for clinical and research applications.