Online native mass spectrometry methods for assaying viral particles
AssigneeRegeneron Pharmaceuticals, Inc.
InventorsVictoria Cotham, Shunhai Wang, Ning Li
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Abstract
Methods for determining the relative abundance of viral capsid components in a sample of viral particles are disclosed. In embodiments, methods for determining the relative abundance of empty capsids, partially-full capsids and full capsids (e.g., containing a heterologous nucleic acid molecule) of adeno-associated virus are disclosed.
Claims20
A method for determining relative abundance of intact viral capsid components in a sample of recombinant viral particles comprising a heterologous nucleic acid molecule, comprising:
- (a) introducing the sample of viral particles into an online native liquid chromatography mass spectrometry (LC-MS) system, wherein the LC-MS system comprises a liquid chromatography column in fluid communication with an electrospray ionization emitter, a mass spectrometer, and a gas inlet port;
- (b) separating the intact viral capsid components in the sample of viral particles via the liquid chromatography column, wherein a mobile phase flow rate through the liquid chromatography column is from 1 μL/min to 20 μL/min;
- (c) contacting the intact viral capsid components with a charge reducing agent via the gas inlet port prior to subjecting the intact viral capsid components to mass spectral analysis;
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- (d) identifying a raw fractional amount of an intact viral capsid component in the sample via mass spectral analysis to determine the relative abundance of each of two or more intact viral capsid components in the sample of viral particles.
A method for determining relative abundance of intact viral capsid components in a sample of recombinant viral particles comprising a heterologous nucleic acid molecule, comprising:
- (a) subjecting the sample of viral particles to online native electrospray ionization mass spectrometry (ESI-MS) to identify a raw fractional amount of an intact viral capsid component, wherein the sample is subjected to chromatographic separation and a charge reducing agent prior to the online native ESI-MS, wherein the chromatographic separation comprises a mobile phase flow rate of from 1 μL/min to 20 μL/min;
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- (b) determining the relative abundance of the intact viral capsid component in the sample of viral particles.
Dependencies are inferred from the claim text.