WO2022062773A1 - 一种小型化、低成本、多衬度无标记显微成像系统 - Google Patents
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Abstract
Description
Claims (9)
- 一种小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述系统包括成像单元,该单元为倒置显微光路结构,包括从上至下依次设置的可编程LED阵列光源、筒镜、微型物镜、样品载物台、彩色相机以及三维位移台;还包括控制单元,用于控制可编程LED阵列光源的照明图案,调节彩色相机的参数,控制相机采集图像,切换成像功能,切换二维/三维成像结果显示,以及执行检测结果分析。
- 根据权利要求1所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述微型物镜的放大倍率为6.4倍,数值孔径NA obj为0.14,焦距4.25,透镜的畸变在1%以内。
- 根据权利要求2所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述可编程LED阵列光源通过改变照明图案可产生六种成像模式照明,包括:明场成像、暗场成像、彩虹暗场成像、莱茵伯格光学染色成像、差分相衬成像以及定量相位成像;所述六种成像模式的照明图案与微型物镜数值孔径相匹配,假设微型物镜数值孔径为NA obj,每个LED对应的照明数值孔径NA ill为:式中,R LED表示每个LED到光轴中心对应的LED的距离,h表示可编程LED阵列光源到样品载物台上表面的距离;明场成像:照明图案为圆形,所有照明的LED的照明数值孔径小于等于微型物镜的数值孔径;暗场成像:照明图案为圆形中空,所有照明的LED的照明数值孔径大于微型物镜的数值孔径;彩虹暗场成像:照明图案与暗场成像的照明图案相同,所有照明的LED的照明数值孔径大于微型物镜的数值孔径,照明光颜色为彩虹分布;莱茵伯格光学染色成像:照明图案为圆形,照明数值孔径小于等于微型物镜数值孔径的LED采用任意颜色,照明数值孔径大于微型物镜数值孔径的LED被不同颜色等分;差分相衬成像:照明图案为半圆形,所有照明的LED的数值孔径小于等于微型物镜的数值孔径;定量相位成像:照明图案为半圆形,所有照明的LED的数值孔径小于等于微型物镜的数值孔径。
- 根据权利要求4所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述定量相位成像通过差分相衬定量相位恢复算法实现,具体过程包括:步骤1,在任意两个轴方向上,沿轴向分别采用两个照明图案照射样品,采集对应的样品图像;假设某一轴向上采集的样品图像分别为I l和I r,另一轴向上采集的样品图像分别为I u和I d,假设采集图像的顺序为I l、I r、I u、I d;步骤2,分别计算两个轴方向上的样品相位梯度图像I lr、I ud:步骤3,根据成像模型,计算差分相衬成像的相位传递函数为:式中,PTF lr(u)表示I l和I r所在轴方向上的相位传递函数,u表示频谱的频率坐标,可被表示为u=(u x,u y),u x和u y为频率横坐标和纵坐标;S lr(u j)表示I l 和I r所在轴方向上的照明光源强度,u j为第j个LED对应的空间频率,P(u)表示光瞳函数,由物镜的数值孔径NA obj和照明波长λ决定,其表现为截止频率为 的圆形滤波函数,P *表示P的共轭;d 2(·)表示二次微分符号;步骤4,求解样品相位:利用Tikhonov准则对差分相衬图像的频谱和相位传递函数进行反卷积计算,获得样品的定量相位结果,计算公式为:式中,φ(r)为样品定量相位,F、F -1分别表示傅里叶变换和傅里叶逆变换,i表示一个轴向的照明,PTF i(u)和PTF i *(u)表示i轴向对应的相位传递函数及其共轭,I i(u)表示i轴向对应的相位梯度图像,β为正则化参数;步骤5,重复上述过程直至停止定量相位成像。
- 根据权利要求5所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述差分相衬定量相位恢复算法,还包括位于步骤4之后的:按照I l、I r、I u、I d的顺序循环采集I l或I r或I u或I d,每采集一幅图像返回执行步骤2。
- 根据权利要求6所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述差分相衬定量相位恢复算法还包括位于步骤5之后的:增强样品定量相位的显示对比度,具体过程包括:直方图滤波:统计相位结果中像素值的灰度分布直方图,通过直方图选择显示的相位值范围将背景相位去除;或者,多颜色伪彩色显示:基于多种伪彩色分布,任意选择相位显示的颜色谱,显示样品的不同结构信息。
- 根据权利要求5所述的小型化、低成本、多衬度无标记显微成像系统,其特征在于,所述相位传递函数生成之后保存至本地,系统在每次启动时,会自动调用相位传递函数。
- 根据权利要求1所述的小型化、低成本、多衬度无标记显微成像系统, 其特征在于,所述成像单元与控制单元之间通过FPGA进行交互,控制单元将控制信息传输至FPGA,由FPGA控制可编程LED阵列光源的照明图案以及控制相机采集图像。
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CN116297337A (zh) * | 2022-09-09 | 2023-06-23 | 山东大学 | 一种使用暗场光学成像技术判别二维材料层数的方法 |
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CN112731641A (zh) * | 2020-12-31 | 2021-04-30 | 安徽医科大学 | 多模式成像手机显微镜装置 |
WO2022241672A1 (zh) * | 2021-05-19 | 2022-11-24 | 浙江大学 | 共聚焦扫描式暗场显微成像方法与装置 |
CN114677447B (zh) * | 2022-03-21 | 2023-05-12 | 中国科学院西安光学精密机械研究所 | 光学显微成像冗余信息定量度量方法 |
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