Multi-Modal Fluorescence Imaging Flow Cytometry System

摘要

In one aspect, the present teachings provide a system for performing cytometry that can be operated in three operational modes. In one operational mode, a fluorescence image of a sample is obtained by exciting one or more fluorophore(s) present in the sample by an excitation beam formed as a superposition of a top-hat-shaped beam with a plurality of beams that are radiofrequency shifted relative to one another. In another operational mode, a sample can be illuminated successively over a time interval by a laser beam at a plurality of excitation frequencies in a scanning fashion. The fluorescence emission from the sample can be detected and analyzed, e.g., to generate a fluorescence image of the sample. In yet another operational mode, the system can be operated to illuminate a plurality of locations of a sample concurrently by a single excitation frequency, which can be generated, e.g., by shifting the central frequency of a laser beam by a radiofrequency. For example, a horizontal extent of the sample can be illuminated by a laser beam at a single excitation frequency. The detected fluorescence radiation can be used to analyze the fluorescence content of the sample, e.g., a cell/particle.

主权项

1. A system for performing cytometry, comprising
a laser for generating a laser beam having a central frequency suitable for exciting at least one fluorophore, an acousto-optic deflector for receiving the laser beam and generating a plurality of angularly separated laser beams including one local oscillator beam (LO beam) and a plurality of RF comb beams, each of said LO beam and the RF comb beams having a radio frequency (RF) shift relative to said central frequency, an optical element for directing said LO beam along a propagation path different than propagation paths of the RF comb beams, a top-hat beam shaper receiving said LO beam and imparting thereto a top-hat intensity profile, a beam splitter receiving said top-hat-shaped LO beam and said RF comb beams and providing a combined excitation beam by spatial overlap of said beams, and at least one optical element for directing said combined beam onto a sample comprising a plurality of cells at least some of which are associated with said fluorophore such that LO beam concurrently illuminates a plurality of spatial locations of the sample and each of said RF comb beams illuminates a different one of said spatial locations to elicit fluorescence radiation from said fluorophore, if present, at said spatial locations, wherein the fluorescence radiation emitted from each of said sample locations exhibits a beat frequency corresponding to a frequency difference between said LO beam and one of the RF comb beams illuminating that sample location.