南方科技大学知识苑(SUSTech KC): 一种高通量集成化磷酸化蛋白组学检测方法

Title	一种高通量集成化磷酸化蛋白组学检测方法
Alternative Title	High-flux integrated phosphorylated proteomics detection method
Author	田瑞军; 李鹏飞; 孔倩
First Inventor	田瑞军
Original applicant	南方科技大学
First applicant	南方科技大学
Address of First applicant	518000 广东省深圳市南山区西丽学苑大道1088号
Current applicant	南方科技大学
Address of Current applicant	518000 广东省深圳市南山区西丽学苑大道1088号 (广东,深圳,南山区)
First Current Applicant	南方科技大学
Address of First Current Applicant	518000 广东省深圳市南山区西丽学苑大道1088号 (广东,深圳,南山区)
Application Number	CN202010426215.5
Application Date	2020-05-19
Open (Notice) Number	CN111579796A
Date Available	2020-08-25
Status of Patent	驳回
Legal Date	2022-10-18
Subtype	发明申请
SUSTech Authorship	First
Abstract	本发明提供了一种高通量集成化磷酸化蛋白组学检测方法，通过将连接在链霉亲和素板上的探针与诱饵蛋白形成捕获骨架，对酪氨酸磷酸化蛋白复合物进行富集捕获，酶解得到多肽，实现减少100倍探针用量的条件下300倍灵敏度的提升。利用150μm内径自填色谱柱高效液相色谱实现短梯度快速分离，结合Q Extractive HF‑X质谱仪在DIA数据采集模式下进行高效检测，可在不损失定性与定量检测性能条件下，实现单个样品35min内分离检测。实现了对不同EGF刺激时间下与不同浓度抑制剂作用下细胞中相关信号通路蛋白磷酸化条件瞬时变化的鉴定。为酪氨酸磷酸化蛋白质复合物的富集与鉴定提供了高通量、高灵敏、快速的检测方法。
Other Abstract	The invention provides a high-flux integrated phosphorylated proteomics detection method. A capture skeleton is formed by a probe connected to a streptavidin plate and bait protein, a tyrosine phosphorylated protein compound is enriched, captured and subjected to enzymolysis to obtain polypeptide, and the sensitivity is improved by 300 times under the condition that the use amount of the probe isreduced by 100 times. Short-gradient rapid separation is realized by utilizing high performance liquid chromatography of a self-filling chromatographic column with the inner diameter of 150 microns; efficient detection is carried out in a DIA data acquisition mode by combining a Q Extractive HF-X mass spectrometer, and separation and detection of a single sample within 35 minutes can be realized under the condition of not losing qualitative and quantitative detection performance. Identification of transient changes of phosphorylation conditions of related signal channel proteins in cells underthe action of inhibitors with different concentrations in different EGF stimulation time is realized. And a high-flux, high-sensitivity and rapid detection method is provided for enrichment and identification of the tyrosine phosphorylated protein compound.
CPC Classification Number	G01N30/02 ; G01N30/72 ; G01N33/6848
IPC Classification Number	G01N33/68 ; G01N30/02 ; G01N30/72
INPADOC Legal Status	（-REJECTION OF INVENTION PATENT APPLICATION AFTER PUBLICATION）[2022-10-18][CN]
INPADOC Patent Family Count	2
Extended Patent Family Count	3
Priority date	2020-05-19
Patent Agent	潘登
Agency	北京品源专利代理有限公司
URL	[Source Record]
Data Source	PatSnap
Document Type	Patent
Identifier	http://kc.sustech.edu.cn/handle/2SGJ60CL/532018
Department	Department of Chemistry
Recommended Citation GB/T 7714	田瑞军,李鹏飞,孔倩. 一种高通量集成化磷酸化蛋白组学检测方法.

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