公开(公告)号：US20210041453A1

公开(公告)日：2021-02-11

申请号：US16979705

申请日：2019-03-12

Applicant: Amgen Inc.

Inventor： Sabrina Amelle BENCHAAR ,  Andrew DYKSTRA

IPC: G01N33/68

Abstract: The disclosed methods are directed to preparing and detecting polypeptides using neutrophil elastase, such as human neutrophil elastase. The polypeptides are optionally denatured, reduced, and/or alkylated before being subjected to a first digestion. A second digestion comprises the use of neutrophil elastase, such as human neutrophil elastase. The prepared fragments are then analyzed chromatographically, electrophoretically, or spectrometrically, or a combination of these methods. The methods are especially useful for the preparation of therapeutic polypeptides for analysis, especially those that are not easily cleaved, such as some bi-specific T-cell engager (BiTE®) molecules.

公开(公告)号：US20230349912A1

公开(公告)日：2023-11-02

申请号：US18026678

申请日：2021-09-17

Applicant: AMGEN INC.

Inventor： Gang XIAO ,  Pavel BONDARENKO ,  Liuqing SHI ,  Thomas M. DILLON ,  Da REN ,  Margaret Speed RICCI ,  Weidong CUI ,  John Joseph HARRAHY ,  Jake PAWLOWSKI ,  Andrew DYKSTRA ,  Dylan RIGGS

IPC: G01N33/68

CPC classification number: G01N33/6821 ,  G01N33/6848 ,  G01N2333/976

Abstract: Provided herein are methods of processing a polypeptide or protein for analysis, e.g., peptide mapping analysis by mass spectrometry. In exemplary embodiments, the method comprises incubating a digested sample at a mildly acidic pH and/or in the presence of a chaotrope, wherein the digested sample is produced by digesting a polypeptide with a protease to produce a digested sample comprising at least two peptides. In exemplary embodiments, the method comprises digesting the polypeptide with a protease which cleaves C-terminal to tryptophan to produce a digested sample comprising at least two peptides. In exemplary embodiments, the method comprises digesting the polypeptide with trypsin at an enzyme:substrate (E:S) weight ratio of about 1:1 to about 1:15 to produce a digested sample comprising at least two peptides. In exemplary aspects, the digested sample comprises at least one or two peptides each comprising a tyrosine at the C-terminus.