Drift-free-high-throughput localization microscopy

Patent metadata

Field	Value
Publication number	US-2026094456-A1
Application number	US-202319113647-A
Country	US
Kind code	A1
Filing date	Oct 30, 2023
Priority date	Oct 31, 2022
Publication date	Apr 2, 2026
Grant date	—

Abstract

A marker-free drift tracking systema and method, referred to as Adaptive localization Intersection based Drift correction (AID), for fast and precise drift correction for single-molecule localization microscopy (SMLM). The system and method utilize an adaptively adjusted position intersection map between two temporally adjacent sets of localized emitter coordinates from the imaging target to achieve robust and precise drift correction at a high computation speed.

First claim

What is claimed is:

1 . A method of processing a plurality of fluorescence image frames obtained from a sample, the method comprising:
receiving a set of localized fluorescent emitter coordinates obtained from the plurality of fluorescence image frames; splitting the set of localized fluorescent emitter coordinates into a series of temporal subsets, wherein each temporal subset is associated with and based on a specified number of the image frames acquired within a certain temporal interval, wherein the temporal subsets include a reference subset, a first drift subset and a plurality of additional drift subsets; creating a first intersected shift-map based on the reference subset and the first drift subset; determining a first peak shift position of the first intersected shift-map; creating a plurality of additional intersected shift-maps using all of the additional drift subsets, wherein each additional intersected shift-map is created based on the reference subset and a respective one of the additional drift subsets; determining an additional peak shift position for each of the additional intersected shift-maps; and determining a drift-corrected localization dataset for the plurality of fluorescence image frames based on the first peak shift position and each of the additional peak shift positions.

2 . The method according to claim 1 , further comprising:
splitting the drift-corrected localization dataset into a series of drift-corrected subsets including a first drift-corrected drift subset and a plurality of additional drift-corrected drift subsets; creating a first drift-corrected intersected shift-map based on the reference subset and the first drift-corrected drift subset; determining a first drift-corrected peak shift position of the first drift-corrected intersected shift-map; creating a plurality of additional drift-corrected intersected shift-maps using all of the additional drift-corrected drift subsets, wherein each additional drift-corrected intersected shift-map is created based on the reference subset and a respective one of the additional drift-corrected drift subsets; determining an additional drift-corrected peak shift position for each of the additional drift-corrected intersected shift-maps; and determining a final drift-corrected localization dataset for the plurality of fluorescence image frames based on the first drift-corrected peak shift position and each of the additional drift-corrected peak shift positions.

3 . The method according to claim 1 , wherein the first intersected shift map is created by calculating a number of the intersected coordinate pairs between the reference subset and a shifted subset created from the first drift subset by shifting coordinates of the drift subset.

4 . The method according to claim 3 , wherein the coordinates of the drift subset are shifted with a step size of D in a local region with a radius of R, where R is larger than a maximum drift between the reference subset and the first drift subset.

5 . The method according to claim 1 , wherein the first peak shift position of the first intersected shift-map is determined from the first intersection shift-map via fast Fourier harmonic analysis.

6 . The method according to claim 1 , the first drift subset and the additional drift subsets having a temporal order, wherein each additional intersected shift-map is created from the reference subset and a respective one of the additional drift subsets in the temporal order of the additional drift subsets.

7 . The method according to claim 6 , wherein a first one of additional intersected shift-maps uses the first peak shift position as its origin, and wherein each remaining ones of the additional intersected shift-maps uses the additional peak shift position determined from an immediately prior additional intersected shift-map as its origin.

8 . The method according to claim 2 , wherein the final drift-corrected localization dataset is determined by estimating time points within each interval separating the temporal subsets using cubic spline interpolation and using the time points to compensate for drift and determine the final drift-corrected localization dataset.

9 . A computer program product including a non-transitory computer readable medium encoded with a computer program product comprising program code for implementing the method of claim 1 .

10 . A microscopy system for obtaining a plurality of fluorescence image frames from a sample, comprising:
at least one light source for illuminating the sample; a positioning stage for holding the sample; at least one detector; and a control system coupled to the at least one light source, the positioning stage and the at least one detector, wherein the control system is structured and configured to:
receive a set of localized fluorescent emitter coordinates obtained from the plurality of fluorescence image frames;
split the set of localized fluorescent emitter coordinates into a series of temporal subsets, wherein each temporal subset is associated with and based on a specified number of the image frames acquired within a certain temporal interval, wherein the temporal subsets include a reference subset, a first drift subset and a plurality of additional drift subsets;
create a first intersected shift-map based on the reference subset and the first drift subset;
determine a first peak shift position of the first intersected shift-map;
create a plurality of additional intersected shift-maps using all of the additional drift subsets, wherein each additional intersected shift-map is created based on the reference subset and a respective one of the additional drift subsets;
determine an additional peak shift position for each of the additional intersected shift-maps; and
determine a drift-corrected localization dataset for the plurality of fluorescence image frames based on the first peak shift position and each of the additional peak shift positions.

11 . The microscopy system according to claim 10 , wherein the first intersected shift map is created by calculating a number of the intersected coordinate pairs between the reference subset and a shifted subset created from the first drift subset by shifting coordinates of the drift subset.

12 . The microscopy system according to claim 11 , wherein the coordinates of the drift subset are shifted with a step size of D in a local region with a radius of R, where R is larger than a maximum drift between the reference subset and the first drift subset.

13 . The microscopy system according to claim 10 , wherein the first peak shift position of the first intersected shift-map is determined from the first intersection shift-map via fast Fourier harmonic analysis.

14 . The microscopy system according to claim 10 , wherein the control system is further structured and configured to:
split the drift-corrected localization dataset into a series of drift-corrected subsets including a first drift-corrected drift subset and a plurality of additional drift-corrected drift subsets; create a first drift-corrected intersected shift-map based on the reference subset and the first drift-corrected drift subset; determine a first drift-corrected peak shift position of the first drift-corrected intersected shift-map; create a plurality of additional drift-corrected intersected shift-maps using all of the additional drift-corrected drift subsets, wherein each additional drift-corrected intersected shift-map is created based on the reference subset and a respective one of the additional drift-corrected drift subsets; determine an additional drift-corrected peak shi

Assignees

• Univ Pittsburgh Commonwealth Sys Higher Education

Inventors

• Ma Hongqiang
• Liu Yang

Classifications

• G06V20/69 Primary

  Microscopic objects, e.g. biological cells or cellular parts · CPC title