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Ultrasonic-based protein quantitation by (18) O-labeling: optimization and comparison between different procedures.

R J Carreira¹, M S Diniz, J L Capelo

¹REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal.

Rapid Communications in Mass Spectrometry : RCM

|December 15, 2010

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Ultrasonic-based direct oxygen-18 labeling significantly accelerates protein quantitation from 12 hours to 15 minutes. This optimized method maintains labeling efficiency, offering a faster alternative for analyzing proteins and complex mixtures.

Area of Science:

Biochemistry
Analytical Chemistry
Proteomics

Background:

Direct oxygen-18 labeling is a method for protein quantitation.
Traditional labeling procedures can be time-consuming, often requiring overnight incubation.

Purpose of the Study:

To optimize and compare ultrasonic-based procedures for direct oxygen-18 labeling.
To accelerate the protein quantitation process using sonication techniques.

Main Methods:

Evaluation of different ultrasonic devices (probe and sonoreactor) and reaction times.
Optimization of enzyme-to-protein ratio and protein concentration.
Application of the optimized method to complex protein mixtures, such as human plasma.

Main Results:

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Sonoreactor-assisted labeling reduced reaction time from 12 hours to 15 minutes without compromising efficiency.
Labeling efficiency showed variability across different peptides, but smaller peptides yielded results comparable to traditional methods.
Sample concentration was identified as a critical factor affecting labeling yield, even in the overnight procedure.

Conclusions:

Ultrasonic-based direct oxygen-18 labeling offers a significantly accelerated and efficient method for protein quantitation.
The optimized protocol is suitable for analyzing both individual proteins and complex biological mixtures.
Further optimization regarding sample concentration may enhance the robustness of the labeling technique.