公开(公告)号：US11639939B2

公开(公告)日：2023-05-02

申请号：US17159136

申请日：2021-01-26

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Yuan Mao ,  Andrew Kleinberg

IPC分类号： G01N33/68 ,  G01N30/72 ,  G01N30/02

摘要： Disclosed are methods of quantifying multiple quality attributes, such as post translational modifications, of multiple samples in a single mass spectrometry (MS) run, including contacting two or more samples with a digesting solution under conditions sufficient to digest samples, wherein each sample is digested separately and the digesting solution is a Tris-free buffer solution; contacting each of the two or more digested samples with a specific Tandem Mass Tag (TMT) labeling reagent under conditions sufficient to label peptides within each of the digested samples with the specific TMT labeling reagent; quenching labeling of peptides within each of the two or more digested samples; combining equal volumes of the two or more labeled, digested samples into a single combined sample solution; and analyzing the single combined sample solution by targeted mass spectral analysis, thereby allowing multiple quality attributes of the two or more samples to be quantified in a single mass spectrometry (MS) run.

公开(公告)号：US20230243841A1

公开(公告)日：2023-08-03

申请号：US17976330

申请日：2022-10-28

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Andrew Kleinberg ,  Yuan Mao ,  Ning Li

IPC分类号： G01N33/68

CPC分类号： G01N33/6848 ,  G01N2560/00

摘要： The present invention generally pertains to methods of preventing disulfide scrambling in non-reducing liquid chromatography-mass spectrometry analysis of a protein of interest. In particular, the present invention pertains to the addition of maleimide to a non-reducing liquid chromatography-mass spectrometry analysis of a protein to prevent disulfide scrambling.

公开(公告)号：US20210231676A1

公开(公告)日：2021-07-29

申请号：US17159136

申请日：2021-01-26

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Yuan Mao ,  Andrew Kleinberg

IPC分类号： G01N33/68 ,  G01N30/72

摘要： Disclosed are methods of quantifying multiple quality attributes, such as post translational modifications, of multiple samples in a single mass spectrometry (MS) run, including contacting two or more samples with a digesting solution under conditions sufficient to digest samples, wherein each sample is digested separately and the digesting solution is a Tris-free buffer solution; contacting each of the two or more digested samples with a specific Tandem Mass Tag (TMT) labeling reagent under conditions sufficient to label peptides within each of the digested samples with the specific TMT labeling reagent; quenching labeling of peptides within each of the two or more digested samples; combining equal volumes of the two or more labeled, digested samples into a single combined sample solution; and analyzing the single combined sample solution by targeted mass spectral analysis, thereby allowing multiple quality attributes of the two or more samples to be quantified in a single mass spectrometry (MS) run.

公开(公告)号：US20220196671A1

公开(公告)日：2022-06-23

申请号：US17554784

申请日：2021-12-17

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Andrew Kleinberg ,  Yuan Mao

IPC分类号： G01N33/68 ,  B01D15/16 ,  B01D15/30 ,  G01N30/72 ,  G01N30/06

摘要： Disclosed are methods for identification of one or more non-native disulfide bonds in a biomolecule (e.g, an antibody). In an example, a method includes performing a digestion of the biomolecule under non-reducing conditions to provide a sample comprising a plurality of biomolecule fragments, contacting the sample to a separation column, applying a first mobile phase gradient comprising trifluoroacetic acid (TFA) and a small molecule additive to the separation column, applying a second mobile phase gradient comprising TFA in acetonitrile (ACN) and a small molecule additive to the separation column, performing a partial reduction procedure on the eluted sample, applying the partially reduced eluted sample components to a mass spectrometer, and performing a mass spectrometric analysis on the partially reduced eluted sample components to identify the one or more non-native disulfide bonds in the biomolecule.

公开(公告)号：US20230349916A1

公开(公告)日：2023-11-02

申请号：US18123753

申请日：2023-03-20

申请人： REGENERON PHARMACEUTICALS, INC.

发明人： Yuan Mao ,  Andrew Kleinberg

IPC分类号： G01N33/68 ,  G01N30/72

CPC分类号： G01N33/6848 ,  G01N30/7233 ,  G01N2030/027

摘要： Disclosed are methods of quantifying multiple quality attributes, such as post translational modifications, of multiple samples in a single mass spectrometry (MS) run, including contacting two or more samples with a digesting solution under conditions sufficient to digest samples, wherein each sample is digested separately and the digesting solution is a Tris-free buffer solution; contacting each of the two or more digested samples with a specific Tandem Mass Tag (TMT) labeling reagent under conditions sufficient to label peptides within each of the digested samples with the specific TMT labeling reagent; quenching labeling of peptides within each of the two or more digested samples; combining equal volumes of the two or more labeled, digested samples into a single combined sample solution; and analyzing the single combined sample solution by targeted mass spectral analysis, thereby allowing multiple quality attributes of the two or more samples to be quantified in a single mass spectrometry (MS) run.

公开(公告)号：US20230417759A1

公开(公告)日：2023-12-28

申请号：US18203854

申请日：2023-05-31

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Yuan Mao ,  Andrew Kleinberg ,  Yunlong Zhao ,  Lili Guo

IPC分类号： G01N33/68 ,  G01N30/34 ,  G01N30/72

CPC分类号： G01N33/6848 ,  G01N33/6857 ,  G01N30/34 ,  G01N30/7233

摘要： A method of enhancing a mass spectral signal is disclosed. The method can include contacting a sample to a separation column under conditions that permit sample components to bind to the substrate; applying a first mobile gradient to the separation column, wherein the first mobile phase gradient comprises trifluoroacetic acid (TFA) and a small molecule additive (e.g., an amino acid) or formic acid (FA) and a small molecule additive (e.g., an amino acid); applying a second mobile gradient to the separation column, wherein the second mobile phase gradient comprises TFA in acetonitrile (ACN) and a small molecule additive (e.g., an amino acid) or formic acid (FA) in ACN and a small molecule additive (e.g., an amino acid); and performing mass spectrometric analysis on eluted sample components.

公开(公告)号：US20210223259A1

公开(公告)日：2021-07-22

申请号：US17143890

申请日：2021-01-07

申请人： Regeneron Pharmaceuticals, Inc.

发明人： Yuan Mao ,  Andrew Kleinberg ,  Yunlong Zhao ,  Lili Guo

IPC分类号： G01N33/68

摘要： A method of enhancing a mass spectral signal is disclosed. The method can include contacting a sample to a separation column under conditions that permit sample components to bind to the substrate; applying a first mobile gradient to the separation column, wherein the first mobile phase gradient comprises trifluoroacetic acid (TFA) and a small molecule additive (e.g., an amino acid) or formic acid (FA) and a small molecule additive (e.g., an amino acid); applying a second mobile gradient to the separation column, wherein the second mobile phase gradient comprises TFA in acetonitrile (ACN) and a small molecule additive (e.g., an amino acid) or formic acid (FA) in ACN and a small molecule additive (e.g., an amino acid); and performing mass spectrometric analysis on eluted sample components.