“光片读出模式”是滨松 sCMOS 相机的独特专利功能,可提高光片显微镜中的信噪比。
光片显微镜利用选择性平面照明,是一种荧光方法,它仅将激发光薄片放置在正在成像的焦点平面上。光片与相机的卷帘快门进行非常精确的同步扫描。卷帘快门的扫描高度有效地变成了狭缝,从而减少了背景信号和散射光。通过使用照明和相机读出的这种组合,可以非常快速地采集光学切片的图像。此外,与许多其他方法相比,样品的光漂白和光毒性显著减少。
光片显微镜用于从小细胞群到整个生物体规模的活细胞成像。在过去几年中,这种技术的使用量急剧增加。
CMOS 图像传感器的曝光和读出方法分为两类:卷帘快门和全局快门。卷帘快门使用连续曝光和读出,将一个像素或一行作为单个单位。因此,在一次图像捕获内曝光定时是不同的。全局快门同时执行所有像素的曝光和读出,一次捕获中的曝光定时是同步的。(参见图 1。)
为了利用此用途中的扫光性质,滨松光子学株式会社使用卷帘快门读出的固有特性和速度开发了光片读出模式。
图 1. 卷帘快门和全局快门的读出定时
sCMOS 相机的标准操作模式是从传感器中心同时从上半部和下半部逐行向外读出每行。(参见图 2。)
使用光片读出模式,用户可以在一次连续扫描中扫描传感器。(参见图 3。)
图 2. 标准模式
图 3. 光片读出模式
光片读出模式提供从上到下和从下到上读出的可选卷帘快门操作。(参见图 4 和图 5。)
图 4. 从上到下读出
图 5. 从下到上读出
光片读出模式中使用的卷帘快门机理允许在下一行曝光开始之前立即重置曝光。此功能通过消除散射光提供更清晰、分辨率更高的图像。
相比之下,使用全局快门,所有像素同时曝光,并且来自照明光片的散射光可能落到相邻行上,从而导致图像模糊。(参见图 6 和图 7。)
图 6. 曝光重置
图 7. 曝光重置的影响
“光片显微镜的柔和照明方案可以前所未有的速度和分辨率对厚样品进行成像。然而,样品内的散射会损害图像质量。现代 sCMOS 相机中的电子共聚焦检测可抑制散射光子,从而获得更好的图像质量。”
Lars Hufnagel 博士
为了使各种激光宽度和扫描速度与传感器同步,光片读出模式提供了灵活的读出间隔,并能够独立设置曝光时间。用户可以轻松地将每行的读出间隔从 10 us 调整到 100 ms。(参见图 8。)
图 8. 可灵活调整读出间隔和曝光时间
外部触发器输出已内置到光片读出模式中,相机可随每行或每帧生成一个外部触发器,使用户能够控制载物台或光源等外围设备。(参见图 9。)
图 9. 外部触发器输出
国家/地区 | 状态 | 专利号 |
---|---|---|
日本 | 已获专利 | JP06475307、JP05770958、JP06240056、JP05639670 |
英国 | 已获专利 | GB2544913、GB2544666、GB2544667、GB2535367、GB2532906、GB2531200、GB2524183、GB2523012、GB2523263、GB2522793 |
德国 | 已获专利 | DE112014006152、DE112014000240、DE112014000195.3 |
美国 | 已获专利 | US10142566、US9423601、US10362245、US10602087 |
中国 | 已获专利 | ZL201480002237.1、ZL201810113454.8、ZL201810113455.2 |
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