公开(公告)号：US20230266214A1

公开(公告)日：2023-08-24

申请号：US17984934

申请日：2022-11-10

Applicant: Wisconsin Alumni Research Foundation

Inventor： Joshua COON ,  Michael WESTPHALL

IPC: G01N1/42 ,  G01N1/36 ,  G01N15/14 ,  H01J49/16 ,  H01J37/28 ,  H01J37/26

CPC classification number: G01N1/42 ,  G01N1/36 ,  G01N15/1468 ,  H01J49/164 ,  H01J49/165 ,  H01J37/28 ,  H01J37/26 ,  H01J2237/31745

Abstract: The present invention provides methods for controllably forming a layer of amorphous ice and other amorphous solids on a substrate, and also provides cryo-electron microscopy (cryo-EM) sample preparation methods and systems that utilize in vacuo formation of amorphous ice and other solids. Formation of the amorphous solid layer can be independent of the deposition of sample molecules to be analyzed using electron microscopy, and allows for the generation of a uniformly thick layer. Optionally, mass spectrometry instruments are used to generate and purify molecules deposited on the generated amorphous solid layer. The techniques and systems described herein can deliver near ideal cryo-EM sample preparation to greatly increase resolution, sensitivity, scope, and throughput of cryo-EM protein imaging, and therefore greatly impact the field of structural biology.

公开(公告)号：US20220065761A1

公开(公告)日：2022-03-03

申请号：US17365698

申请日：2021-07-01

Applicant: Wisconsin Alumni Research Foundation

Inventor： Joshua COON ,  Michael WESTPHALL

IPC: G01N1/42 ,  G01N1/36 ,  G01N15/14 ,  H01J49/16 ,  H01J37/28 ,  H01J37/26

Abstract: The present invention provides methods for controllably forming a layer of amorphous ice and other amorphous solids on a substrate, and also provides cryo-electron microscopy (cryo-EM) sample preparation methods and systems that utilize in vacuo formation of amorphous ice and other solids. Formation of the amorphous solid layer can be independent of the deposition of sample molecules to be analyzed using electron microscopy, and allows for the generation of a uniformly thick layer. Optionally, mass spectrometry instruments are used to generate and purify molecules deposited on the generated amorphous solid layer. The techniques and systems described herein can deliver near ideal cryo-EM sample preparation to greatly increase resolution, sensitivity, scope, and throughput of cryo-EM protein imaging, and therefore greatly impact the field of structural biology.

公开(公告)号：US20180286649A1

公开(公告)日：2018-10-04

申请号：US15936288

申请日：2018-03-26

Applicant: Wisconsin Alumni Research Foundation

Inventor： Joshua COON ,  Nicholas RILEY ,  Michael WESTPHALL

IPC: H01J49/00 ,  H01J49/42 ,  G01N33/68

Abstract: A new approach is described herein for outfitting a mass spectrometer with an infrared laser that provides an improved method of ion dissociation. One embodiment, generally referred to as Activated Ion Electron Transfer Dissociation (AI-ETD) utilizes additional energy from photons during fragmentation to generate extensive fragmentation by interacting with peptides or proteins that are not fully fragmented or separated in the high pressure linear ion trap, thus allowing for increased information during MS/MS. Additionally, a new activation scheme generally referred to as AI-ETD+ is also described that combines AI-ETD in the high pressure cell of the linear ion trap with additional infrared multi-photon dissociation (IRMPD) activation in the low pressure cell. These methods provide improved fragmentation and sequence coverage without introducing additional time to the scan duty cycle.

公开(公告)号：US20240060863A1

公开(公告)日：2024-02-22

申请号：US18261267

申请日：2022-01-13

Applicant: Wisconsin Alumni Research Foundation

Inventor： Michael WESTPHALL ,  Joshua COON

IPC: G01N1/42 ,  G01N1/28

CPC classification number: G01N1/42 ,  G01N1/2813

Abstract: The present invention provides an improved technique for cryogenically fixing biological samples in amorphous ice for analysis by cryo-electron microscopy (cryo-EM). Analyte particles are cooled to very low temperatures prior to depositing the particles onto a cooled substrate surface, such as a transmission electron microscope (TEM) grid. This approach “locks” in the particle structure prior to deposition. Either concurrently with or after deposition, the analyte particles are further contacted with a vapor stream of atoms or molecules at cryogenic or near cryogenic temperatures. As a result, a thin layer of an amorphous solid is formed around each particle without significant conformational changes in the particle structure, thereby forming an improved sample for EM analysis.

公开(公告)号：US20200158607A1

公开(公告)日：2020-05-21

申请号：US16626250

申请日：2018-07-06

Applicant: Wisconsin Alumni Research Foundation

Inventor： Joshua COON ,  Michael WESTPHALL

IPC: G01N1/42 ,  H01J49/16 ,  G01N1/36 ,  G01N15/14

Abstract: The present invention provides methods for controllably forming a layer of amorphous ice and other amorphous solids on a substrate, and also provides cryoelectron microscopy (cryo-EM) sample preparation methods and systems that utilize in vacuo formation of amorphous ice and other solids. Formation of the amorphous solid layer can be independent of the deposition of sample molecules to be analyzed using electron microscopy, and allows for the generation of a uniformly thick layer. Optionally, mass spectrometry instruments are used to generate and purify molecules deposited on the generated amorphous solid layer. The techniques and systems described herein can deliver near ideal cryo-EM sample preparation to greatly increase resolution, sensitivity, scope, and throughput of cryo-EM protein imaging, and therefore greatly impact the field of structural biology.