Development and Application of an MRM Method for Simultaneous Quantification of Sodium Channels Na v 1.1, Na v 1.2 and Na v 1.6 in Stable HEK293 cell lines, and Solubilized Membrane Proteins from Rodents and Human Brain Tissues.

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Abstract
Rationale Na v 1.1, 1.2 and 1.6 are transmembrane proteins acting as voltage gated sodium channels implicated in various forms of epilepsy. There is a need for knowing their actual concentration in target tissues during drug development. Methods Unique peptides for Na v 1.1, Na v 1.2 and Na v 1.6 were selected as quantotropic peptides for each protein and used for their quantification in plasma membranes from stably transfected HEK293 cells and rodent and human brain samples using ultra‐high‐performance liquid chromatography/electrospray ionization tandem mass spectrometry. Results Na v 1.1, 1.2 and 1.6 protein expressions in three stably individually transfected HEK293 cell lines were found to be 2.1±0.2, 6.4±1.2 and 4.0±0.6 fmole/µg membrane protein respectively. Na v 1.2 showed the highest expression, with approximately 3 times higher (p> Na v 1.1> Na v 1.6, with the human brain expressing much lower concentrations overall in comparison to rodents.