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多功能高灵敏度发光分析仪(LUANA):用于凝胶电泳

Multipurpose high sensitivity luminescence analyzer (LUANA): use in gel electrophoresis.

作者信息

Neri D, Prospero T, Petrul H, Winter G, Browne M, Vanderpant L

机构信息

Cambridge Center for Protein Engineering-MRC Centre, England, UK.

出版信息

Biotechniques. 1996 Apr;20(4):708-13.

PMID:8800693
Abstract

Many applications in molecular biology require the rapid and high-sensitivity detection of biological macromolecules. Here we describe a luminescence analyzer (LUANA), featuring xenon lamp-based illumination and a cooled CCD camera as detector. Luminescent samples (gels, membranes or microplates) are placed in a light-tight chamber, and computer software is used to control the camera and to display and analyze the images. LUANA can be used with a broad spectrum of excitation and emission wavelengths (200-1000 nm), allowing a choice of many fluorescent dyes and facilitating multicolor imaging. We show that LUANA is readily applicable to gel electrophoresis and highly sensitive; on gels we could detect < 100 fg of Cy5TM-labeled protein and 0.1 ng of plasmid DNA labeled with a fluorescent intercalator. We observed a linear response for sample concentrations differing by two orders of magnitude using a single acquisition time. This range could be further extended by using different acquisition times. We have also used LUANA in two-color imaging of DNA and for screening of antigen-antibody and peptide-protein complexes by gel bandshift.

摘要

分子生物学中的许多应用都需要对生物大分子进行快速且高灵敏度的检测。在此,我们描述一种发光分析仪(LUANA),其特点是以氙灯为照明光源,并配备冷却式电荷耦合器件(CCD)相机作为探测器。将发光样本(凝胶、膜或微孔板)置于不透光的腔室中,利用计算机软件控制相机并显示和分析图像。LUANA可在宽光谱的激发和发射波长范围(200 - 1000纳米)内使用,这使得可以选择多种荧光染料并便于进行多色成像。我们证明LUANA很容易应用于凝胶电泳且灵敏度高;在凝胶上,我们能够检测到<100飞克的Cy5TM标记蛋白以及0.1纳克用荧光嵌入剂标记的质粒DNA。使用单一采集时间,我们观察到样本浓度相差两个数量级时呈现线性响应。通过使用不同的采集时间,这个范围还可以进一步扩大。我们还将LUANA用于DNA的双色成像以及通过凝胶带迁移筛选抗原 - 抗体和肽 - 蛋白质复合物。

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