Absolute Quantification of Nav1.5 Expression by Targeted Mass Spectrometry

Area of Science:

• Cardiovascular Biology
• Proteomics
• Biomarker Discovery

Background:

• Nav1.5 is crucial for cardiac action potential and heartbeat regulation.
• Abnormal Nav1.5 levels are linked to cardiac diseases like Brugada syndrome.
• Existing methods only measure relative Nav1.5 quantities, posing diagnostic challenges.

Purpose of the Study:

• To develop the first targeted, mass spectrometry-based assay for absolute quantification of Nav1.5 protein.
• To establish a reliable method for measuring Nav1.5 copy numbers in biological samples.
• To overcome the challenges in quantifying large, transmembrane proteins like Nav1.5.

Main Methods:

• Development of a targeted mass spectrometry assay.
• Quantification of Nav1.5 in Chinese hamster ovary (CHO) cells.
• Determination of the lower limit of quantification (LLOQ) and precision.

Main Results:

• The first absolute quantification of Nav1.5 protein in a biological sample.
• Quantification of transiently expressed Nav1.5 in 1.5 million CHO cells.
• Achieved a sensitivity of 3 ± 2 × 10^4 copies/cell.

Conclusions:

• The developed assay provides absolute Nav1.5 quantification, a significant advancement for cardiac research.
• This method enables precise measurement of Nav1.5 levels, crucial for diagnosing and treating related cardiac conditions.
• This is the first absolute quantitation of Nav1.5 in any biological sample, setting a new standard for protein analysis.