Computerized analysis of double‐label autoradiographs: A screening method for 1,25‐dihydroxyvitamin D3 induced changes in intestinal protein synthesis
Computerized analysis of double‐label autoradiographs: A screening method for 1,25‐dihydroxyvitamin D3 induced changes in intestinal protein synthesis
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双标记放射自显影图的计算机分析：1,25-二羟基维生素 D3 诱导的肠道蛋白质合成变化的筛选方法
DOI：
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发表时间：
1982
期刊：
影响因子：
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通讯作者：
H. DeLuca
中科院分区：
文献类型：
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作者：
N. Kendrick;K. Eakle;H. DeLuca
威斯康星大学的McIDAS计算机系统(人机交互数据访问系统)被用来分析二维(2-D)电泳胶的双标记放射自显影。用来产生2-D凝胶的4个样品取自8只维生素D缺乏的大鼠肠道，注射1，25-(OH)2D3(1，25-(OH)2D3)或赋形剂后25h。为了进行分析，将柯达XAR5(XR)胶片和柯达NS-2T(NS)胶片上的图案转换为数字图像，柯达XAR5(XR)胶片在-75°C下通过荧光照相检测14C+3H，而柯达NS-2T(NS)胶片在室温下仅通过直接放射自显影检测14C。使用McIDAS系统将NS图像乘以归一化系数，该归一化系数使其与XR图像的曝光相匹配。从两幅图像中减去背景后，将XR图像除以对齐的归一化NS图像；将得到的比率图像乘以增强因子，并根据像素亮度进行着色。相对于平均比例高或低比例的多肽在比例图像上的对比色中明显突出，可以通过肉眼检测到。然后可以通过斑点积分来量化这些高亮斑点的实际XR/NS密度比。为了测试该方法，测量了4个胶片上每一个上的34个点的XR/NS密度比率，以及与比率图像上的斑点外观相比较的斑点比率。除了几个假阳性外，一致意见都很好。几种多肽的相对平均比例增加了25-50%，而一种相对分子质量为50000的多肽，约等电点5.5，平均增加了240%。
The McIDAS computer system (Man‐computer Interactive Data Access System) at the University of Wisconsin was used to analyze double‐label autoradiographs of two‐dimensional (2‐D) electrophoresis gels. The 4 samples used to generate the 2‐D gels were obtained from 8 vitamin D‐deficient rat intestines 25 h after injection with 1,25‐dhydroxyvitamin D3 (1,25‐(OH)2 D3) or vehicle. To perform the analysis, the patterns on Kodak XAR5 (XR) film, which detects 14C + 3H by fluorography at ‐75 °C, and on Kodak NS‐2T (NS) film, which detects 14C only by direct autoradiography at room temperature, were converted to digital images. The McIDAS system was used to multiply the NS image by a normalization factor which matched it to the XR image with respect to exposure. After background subtraction from both images, the XR image was divided by the aligned normalized NS image; the resultant ratio image was multiplied by an enhancement factor and colored according to pixel brightness. Polypeptides with a high or low ratio relative to the average ratio stood out clearly in contrasting colors on the ratio image and could be detected by visual inspection. The actual XR/NS density ratios of these highlighted spots could then be quantitated by spot integration. To test the method, XR/NS density ratios were measured for 34 spots on each of the 4 films, sets and the spot ratios compared to the spot appearances on the ratio image. With the exception of a few false positives, agreement was good. Several polypeptides were found to be increased by 25–50 % relative to the average ratio while one polypeptide of molecular weight 50 000, approximate pI 5.5, was increased an average of 240 %.
维生素 D 缺乏对鸡肠道蛋白体外标记的影响：二维电泳分析。
影响因子：
2.9
作者：
Kendrick,NC;Barr,CR;Moriarity,D;DeLuca,HF
通讯作者：
DeLuca,HF