Microarray analysis in clinical oncology: pre-clinical optimization using needle core biopsies from xenograft tumors.
Microarray analysis in clinical oncology: pre-clinical optimization using needle core biopsies from xenograft tumors.
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DOI：
10.1186/1471-2407-4-20
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发表时间：
2004-05-19
期刊：
影响因子：
3.8
通讯作者：
Camphausen K
中科院分区：
文献类型：
--
作者：
Goley EM;Anderson SJ;Ménard C;Chuang E;Lü X;Tofilon PJ;Camphausen K
对临床组织样本进行DNA微阵列分析可能会提供有关人类癌症生物学的重要信息。然而，活检芯是人类肿瘤组织的典型来源，通常提供非常少量的核糖核酸(0.3-15μg)。RNA扩增是一种常见的方法，用来增加杂交实验可用的材料数量。利用人类异种移植组织，我们试图解决以下三个问题：1)扩增的RNA是否代表原始的RNA图谱？2)进行代表性扩增所需的总RNA的最小数量是多少？3)标记杂交探针的直接方法和间接方法是否相同？从人的异种移植组织中提取总RNA，通过线性扩增过程进行扩增。RNA被标记和杂交，产生的图像产生的数据被提取到两类使用mAdb系统：“所有基因”和“离群值”。每张幻灯片都生成了散点图，并获得了皮尔逊相关系数。结果表明，扩增5μg的总RNA样品与总RNA样品之间的皮尔森相关系数为0.752(N=6,987个基因)。我们随后确定，与相应的原始RNA样本相比，扩增0.5RNAg产生了类似的皮尔森相关系数。同样，扩增5μg的起始核糖核酸可检测到69%的总核糖核酸异常值，0.5μg的起始核糖核酸可检测到55%的异常值。然而，扩增0.05μg的起始RNA会导致保真度的损失(扩增的样品和原始样品之间的皮尔逊系数为0.669，44%的异常值一致性)。在这些研究中，直接或间接的探针标记方法产生了类似的结果。最后，我们检查了从人肿瘤异种移植的针芯活检中获得的RNA，扩增并间接标记后，与更大的切除活检材料相比，是否会产生典型的阵列特征。在此分析中，U251活检点与切除肿瘤的相关系数为0.750-0.834，DU145活检点与切除肿瘤的相关系数为0.812-0.846。这些数据表明，经过线性扩增和起始RNA的间接或直接标记后，针芯活检可以作为可靠的组织样本用于肿瘤微阵列分析。
DNA microarray profiling performed on clinical tissue specimens can potentially provide significant information regarding human cancer biology. Biopsy cores, the typical source of human tumor tissue, however, generally provide very small amounts of RNA (0.3–15 μg). RNA amplification is a common method used to increase the amount of material available for hybridization experiments. Using human xenograft tissue, we sought to address the following three questions: 1) is amplified RNA representative of the original RNA profile? 2) what is the minimum amount of total RNA required to perform a representative amplification? 3) are the direct and indirect methods of labeling the hybridization probe equivalent? Total RNA was extracted from human xenograft tissue and amplified using a linear amplification process. RNA was labeled and hybridized, and the resulting images yielded data that was extracted into two categories using the mAdb system: "all genes" and "outliers". Scatter plots were generated for each slide and Pearson Coefficients of correlation were obtained. Results show that the amplification of 5 μg of total RNA yields a Pearson Correlation Coefficient of 0.752 (N = 6,987 genes) between the amplified and total RNA samples. We subsequently determined that amplification of 0.5 μg of total RNA generated a similar Pearson Correlation Coefficient as compared to the corresponding original RNA sample. Similarly, sixty-nine percent of total RNA outliers were detected with 5 μg of amplified starting RNA, and 55% of outliers were detected with 0.5 μg of starting RNA. However, amplification of 0.05 μg of starting RNA resulted in a loss of fidelity (Pearson Coefficient 0.669 between amplified and original samples, 44% outlier concordance). In these studies the direct or indirect methods of probe labeling yielded similar results. Finally, we examined whether RNA obtained from needle core biopsies of human tumor xenografts, amplified and indirectly labeled, would generate representative array profiles compared to larger excisional biopsy material. In this analysis correlation coefficients were obtained ranging from 0.750–0.834 between U251 biopsy cores and excised tumors, and 0.812–0.846 between DU145 biopsy cores and excised tumors. These data suggest that needle core biopsies can be used as reliable tissue samples for tumor microarray analysis after linear amplification and either indirect or direct labeling of the starting RNA.
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影响因子：
14.9
作者：
Baugh, L. R.;Hill, A. A.;Hunter, Craig P.
通讯作者：
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