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Semrock 滤光片

Semrock 滤光片

FISH(荧光免疫原位杂交)

荧光原位杂交用于通过互补探针序列的杂交来显示确定的核酸序列。FISH 有很多应用,从基本的 DNA 定位到染色体畸变的诊断(细胞遗传学)。多色 FISH 图像分析需要使用单独的滤光片激发块(立方体)或激发滤光片转轮结合多波段二向色镜和发射滤光片来区别各种信号。

Crosstalk value chart per fluorophores filter set

荧光免疫原位杂交(FISH)和 多路复用(Densely Multiplexed)成像的串色

当同时使用具有多个密集光谱的荧光染料时,区分荧光标记的能力决定了检测结果的速度和准确性。在进行荧光免疫原位杂交(FISH)测量时,这种密集的图像复用特别重要。因此,减少串色(即来自不希望的荧光染料的信号相对于所需荧光染料的信号影响)至关重要。下表量化了使用特定的 BrightLine 荧光免疫原位杂交(FISH)滤光片组时,相邻荧光染料之间的串扰值。该串扰值决定于典型的归一化荧光染料光谱、滤光片的设计光谱和强金属卤化物灯光谱的重叠。

SpectrumRed filter set

下图显示了滤光片组的光谱曲线(蓝色、绿色和红色实线)和 荧光染料 SpectrumGold™、SpectrumRed™ 和 Cy5™(从左到右)的吸收光谱和发射光谱曲线。 

  Crosstalk value chart per fluorophores filter set

例如,当使用 SpectrumGold™ 滤光片组对标记有 SpectrumGreen™、SpectrumGold™ 和 SpectrumRed™ 荧光染料的样品进行成像时,不需要的 SpectrumGreen 信号将小于所需要的 SpectrumGold 信号的 2%,并且 SpectrumRed 信号将小于1%。

好的滤光片会带来哪些不同 ?

这些 BrightLine 滤光片组经过精心优化,能显著增加常用荧光染料的亮度,同时降低不必要的背景噪声的强度,也能减少与相邻荧光染料的串扰。如图所示:各荧光通道之间的信号串扰(串色)只有几个百分点或更少。以上图表显示了定量比较。

使用软镀膜滤光片组(左)和 BrightLine 滤光片组(右)

human tumor cell image viewed with Semrock Brightline FISH filter versus competitor

人类肿瘤与 CEP 3 探针杂交,Spectrum Red 结合了 CEP 探针(为 Vysis-Urovysion™ 分析的一部分,放大400X),等量曝光时间对照比较。图片来源:Tina Bocker Edmonston,MD,Thomas Jefferson University

 

integration time graph of Semrock Brightline FISH filter set versus competitor's filters

适用于查找中期相等自动化任务。这个行业测试比较了 BrightLine FISH 滤光片组和软镀膜滤光片组所需的积分时间。该自动化系统基于蔡司 AXIO 成像显微镜,使用相同的图像强度来找到中期相。BrightLine FISH 滤光片组的积分时间更短,成像速度更快。

了解有关 FISH 的更多信息:

技术说明

荧光滤光片简介

装配滤光片镜盒

多带滤光片组配置术语

相关 出版物

获取荧光图像的联合 – 2005年8月刊  BioPhotonics

新型滤光片改变了高速多色荧光成像 2006 年3月刊 BioPhotonics

更多资源

尼康显微镜 U – 荧光原位杂交、研究实验室中的硬件和软件意义

 

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