Targeted and Untargeted Amine Metabolite Quantitation in Single Cells with Isobaric Multiplexing

Area of Science:

• Biochemistry
• Analytical Chemistry
• Cell Biology

Background:

• Accurate quantification of intracellular molecules is crucial for understanding cellular function.
• Previous methods lacked the sensitivity and throughput for single-cell amine analysis.

Purpose of the Study:

• To develop and validate a sensitive, high-throughput method for single-cell amine and amino acid quantification.
• To enable absolute quantification of intracellular amino acids and relative quantification of amines in individual cells.

Main Methods:

• Isobaric multiplexing of 6 individual cells with an analyte-abundant carrier.
• Nanoflow liquid chromatography coupled with high-resolution mass spectrometry (LC-MS).
• Targeted quantification of 10 amino acids and untargeted analysis of other amines.

Main Results:

• Achieved low attomole limits of detection and linear calibration ranges for targeted amino acids.
• Determined millimolar intracellular amino acid concentrations using cell size data.
• Identified 249 amine features with modest cell type separation via principal component analysis.

Conclusions:

• The developed method offers accessible, sensitive, and high-throughput single-cell amine analysis.
• This approach facilitates precise intracellular concentration measurements.
• The methodology holds potential for expansion to other ultrasensitive analyses.