SCANNING PARTIAL CONFOCAL MICROSCOPE

扫描局部共焦显微镜

• 批准号: 2285121
• 负责人: WALTER A. CARRINGTON
• 金额: $ 15.5万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-01-01 至 1997-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2285121
• 关键词: animal tissue; biomedical equipment development; charge coupled device camera; computer program /software; computer system design /evaluation; confocal scanning microscopy; fluorescent dye /probe; mathematical model; model design /development

A conventional epifluorescence microscope exhibits a partial confocal effect when a small illumination aperture is used. This effect causes a reduction in the out of focus light present when viewing a 3D object. Inserting smaller apertures than the usual iris diaphragm causes a large confocal effect; an illumination spot at the sample of, for example, 2 microns across results in the nearly complete absence of out of focus light originating more than 2 microns out of focus. We propose to develop instruments based on this effect combined with computational methods called image restoration or deconvolution that reverse the blurring of the optics and remove any remaining out of focus light. We will notify two conventional epifluorescence microscopes, one which will scan the sample across the illumination spot and another that will scan the illumination spot across the sample. Only the illumination path is changed by replacing the usual iris diaphragm by a smaller aperture. The light collection path is entirely unchanged, so high quantum efficacy CCD cameras can still be used; thus the light collection loses of conventional confocal microscopes are avoided. These instruments will have significant advantages for imaging of fluorescently labeled biological samples. They will provide high resolution 3D images in thick samples where there is too much out of focus light for seeing fine detail in conventional microscopes. In combination with image restoration these instruments will provide greater resolution, contrast, and accuracy of quantitative fluorescence measurements in living fluorescently labelled samples that is possible with either conventional side field or conventional confocal microscopes. This new instrument will be tested and applied to studies directed at understanding the control of vascular function.

常规的落荧光显微镜表现出部分共共共聚焦 当使用小照明光圈时的效果。 这种效果会导致 查看3D对象时，会减少焦点灯的降低。 插入比通常的虹膜隔膜的小光圈会导致大的孔径 共聚焦效应；例如2 在几乎完全没有焦点的情况下，整个结果的微米 光源超过2微米的光不聚焦。 我们建议 根据这种效果与计算相结合开发仪器 称为图像恢复或反卷积的方法可以逆转 光学元件的模糊，并删除剩余的任何从聚焦光中。 我们 将通知两个常规的落荧光显微镜，一个将 扫描样品穿过照明点，另一个将扫描 样品中的照明位置。 只有照明路径是 通过用较小的光圈代替通常的虹膜膜片来更改。 这 光收集路径完全没有变化，因此高量子功效CCD 仍然可以使用相机；因此，光收集失去了 避免常规共聚焦显微镜。 这些乐器会 具有荧光标记的成像具有显着优势 生物样品。 他们将提供高分辨率的3D图像 样品中有太多聚焦光线无法查看细节的样本 在常规显微镜中。 结合图像恢复这些 仪器将提供更大的分辨率，对比度和准确性 活荧光标记的定量荧光测量 常规侧面字段或 常规共聚焦显微镜。 该新仪器将进行测试 并应用于旨在了解血管控制的研究 功能。