Flat field correction technique for time delay integration fluorescence microscopy imaging

Songtao CHANG; Haojie XIA

doi:10.37188/OPE.20223005.0527

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Flat field correction technique for time delay integration fluorescence microscopy imaging

Modern Applied Optics | 更新时间：2022-03-25

- Flat field correction technique for time delay integration fluorescence microscopy imaging
- Optics and Precision Engineering Vol. 30, Issue 5, Pages: 527-535(2022)
- 作者机构：
  
  1.合肥工业大学仪器科学与光电工程学院，安徽合肥230009
  2.测量理论与精密仪器安徽省重点实验室，安徽合肥230009
- 作者简介：
- 基金信息：
- DOI：10.37188/OPE.20223005.0527
  CLC： TH742;R318.6
- Received：04 August 2021，
  
  Revised：22 September 2020，
  
  Published：10 March 2022
- 稿件说明：
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常松涛,夏豪杰.时间延迟积分荧光显微成像平场校正技术[J].光学精密工程,2022,30(05):527-535.

CHANG Songtao,XIA Haojie.Flat field correction technique for time delay integration fluorescence microscopy imaging[J].Optics and Precision Engineering,2022,30(05):527-535.
常松涛,夏豪杰.时间延迟积分荧光显微成像平场校正技术[J].光学精密工程,2022,30(05):527-535. DOI： 10.37188/OPE.20223005.0527.

CHANG Songtao,XIA Haojie.Flat field correction technique for time delay integration fluorescence microscopy imaging[J].Optics and Precision Engineering,2022,30(05):527-535. DOI： 10.37188/OPE.20223005.0527.

摘要

时间延迟积分（Time Delay Integration， TDI）图像传感器具有高速、高灵敏度等特点，广泛应用于高通量、大视场的荧光显微成像系统中。显微物镜视场内响应均匀是精确获取荧光能量分布的基础，为提高系统成像质量和测量准确度，研究了适用于TDI荧光显微成像系统的平场校正或响应非均匀性校正方法。根据TDI荧光成像系统的工作原理推导激光诱导荧光成像模型，分析图像均匀性退化机理。提出一种基于微阵列生物芯片的两步式校正方法，将系统响应非均匀性分为高频和低频部分分别校正，高频部分采用传统的两点校正方法，低频部分采用微阵列生物芯片校正。基于高通量TDI荧光显微成像系统开展实验，执行并验证本文的校正方法。实验结果表明：本文方法将TDI荧光成像系统的响应非均匀性由25.21%降低至2.87%，显著提高了系统性能。本文提出的校正方法能够满足TDI荧光显微成像系统的平场校正需求，具有一定的应用价值。

Abstract

Owing to their high frame rate and sensitivity， time delay integration （TDI） image sensors are widely used in fluorescence microscopy imaging systems with high throughput and large field of view. Uniformity of response in the field of view of the microscope objective is the basis for accurate acquisition of fluorescence energy distribution. To improve the imaging quality and measurement accuracy of the system， this study investigated the response non-uniformity correction method applicable to the TDI fluorescence microscopy imaging system. First， a laser-induced fluorescence imaging model was devised based on the working principle of the TDI fluorescence imaging system， and the mechanism of image uniformity degradation was analyzed. A two-step correction method based on microarray biochips was then proposed， which divided the nonuniformity of system response into high frequency and low-frequency parts to be corrected separately. The former can be corrected by traditional two point correction， whereas the latter is corrected using the proposed microarray biochip-based method. Finally， experiments based on a high-throughput TDI fluorescence microscopy imaging system were conducted to verify the calibration method employed in this study. The experimental results show that the proposed approach reduces the response nonuniformity of the TDI fluorescence imaging system from 25.21% to 2.87%， implying a significant improvement in the system performance. Moreover， it shows that the correction method proposed in this paper can significantly correct the response nonuniformity of TDI fluorescent imaging systems， and is therefore effective and practical.

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