Quantification of Low Copy Number Proteins in Single Cells

Area of Science:

• Biochemistry
• Analytical Chemistry
• Cell Biology

Background:

• Quantifying low copy number intracellular proteins in single cells is challenging.
• Existing methods often lack the sensitivity or selectivity required for such analyses.

Purpose of the Study:

• To develop an ultrasensitive and highly selective method for quantifying low copy number intracellular proteins in single cells.
• To demonstrate the method's capability by measuring active caspase3 protein levels in Jurkat cells.

Main Methods:

• Utilized a low-cost laser-induced fluorescence (LIF) detector and a BV605 fluorescent probe.
• Employed capillary electrophoresis (CE)-LIF analysis with in situ lysis for single-cell protein quantification.
• Separated and distinguished proteins based on retention time differences.

Main Results:

• Achieved detection of approximately seven active caspase3 protein molecules in a 91 pL detection volume.
• Quantified active caspase3 molecules in 98 single Jurkat cells, ranging from 629 to 12171 molecules.
• Demonstrated significant heterogeneity in protein levels among cells from the same batch.

Conclusions:

• The developed CE-LIF method offers high sensitivity and selectivity for single-cell protein quantification.
• The technique can be extended to quantify other low copy number proteins with appropriate antibodies.
• This low-cost system holds promise for early cancer diagnosis, disease pathway research, and high-throughput applications.