Comparison of frozen and RNALater solid tissue storage methods for use in RNA expression microarrays.
Comparison of frozen and RNALater solid tissue storage methods for use in RNA expression microarrays.
复制标题
DOI：
10.1186/1471-2164-5-88
复制
发表时间：
2004-11-10
期刊：
影响因子：
4.4
通讯作者：
Warrington JA
中科院分区：
文献类型：
--
作者：
Mutter GL;Zahrieh D;Liu C;Neuberg D;Finkelstein D;Baker HE;Warrington JA
原代人体组织是发现表征疾病状态的基因表达模式的无价的广泛使用的工具。组织处理方法仍然不标准化，导致如何最好地储存收集的组织和保持实验室之间的再现性的问题没有得到解决。我们细分子宫肌层组织标本，并使用最常见的组织处理方法（新鲜、冷冻、RNALater）储存等分试样，然后比较Affytek U133人类表达阵列上的定量RNA表达谱。在每个处理组中分割样本并纳入重复样本，使我们能够进行正式的全基因组分析，比较样本来源（不同患者）、处理方案（新鲜vs.冷冻vs. 24或72小时RNALater）和随机背景（重复样本）导致的结果变异程度。对数据集进行随机排列，以定义ANOVA检验统计值的基线模式，并据此解释观察到的结果。22,283个基因中有14,639个在至少一个样本中表达。患者受试者在混合模型ANOVA中提供了最大的变异来源，重复和处理方法最少。处理方法（24小时RNALater vs 72小时RNALater vs新鲜vs冷冻）产生的变异程度与重复测定中观察到的变异性相似。根据基因功能分类的检验统计量的子集分析表明，每个功能分类内的“离群值”ANOVA结果的频率总体上不大于偶然的预期。相对于新鲜或冷冻组织的替代物，在RNALater中室温储存组织24或72小时不会导致定量RNA表达结果的任何系统性变化。这种无毒的防腐剂使得能够分散组织收集用于表达阵列分析，而不需要专门的设备。
Primary human tissues are an invaluable widely used tool for discovery of gene expression patterns which characterize disease states. Tissue processing methods remain unstandardized, leading to unanswered concerns of how to best store collected tissues and maintain reproducibility between laboratories. We subdivided uterine myometrial tissue specimens and stored split aliquots using the most common tissue processing methods (fresh, frozen, RNALater) before comparing quantitative RNA expression profiles on the Affymetrix U133 human expression array. Split samples and inclusion of duplicates within each processing group allowed us to undertake a formal genome-wide analysis comparing the magnitude of result variation contributed by sample source (different patients), processing protocol (fresh vs. frozen vs. 24 or 72 hours RNALater), and random background (duplicates). The dataset was randomly permuted to define a baseline pattern of ANOVA test statistic values against which the observed results could be interpreted. 14,639 of 22,283 genes were expressed in at least one sample. Patient subjects provided the greatest sources of variation in the mixed model ANOVA, with replicates and processing method the least. The magnitude of variation conferred by processing method (24 hours RNALater vs 72 hours RNALater vs. fresh vs frozen) was similar to the variability seen within replicates. Subset analysis of the test statistic according to gene functional class showed that the frequency of "outlier" ANOVA results within each functional class is overall no greater than expected by chance. Ambient storage of tissues for 24 or 72 hours in RNALater did not contribute any systematic shift in quantitative RNA expression results relative to the alternatives of fresh or frozen tissue. This nontoxic preservative enables decentralized tissue collection for expression array analysis without a requirement for specialized equipment.
登录
查看更多内容
DOI：
10.1016/0005-2744(74)90240-x
发表时间：
1974-01-01
期刊：
BIOCHIMICA ET BIOPHYSICA ACTA
影响因子：
--
作者：
ALLEWELL, NM;SAMA, A
通讯作者：
SAMA, A
影响因子：
2.1
作者：
Bachoon, DS;Chen, F;Hodson, RE
通讯作者：
Hodson, RE
影响因子：
2.7
作者：
Barrett, MT;Glogovac, J;Rabinovitch, PS
通讯作者：
Rabinovitch, PS
影响因子：
2.2
作者：
Medeiros, M;Sharma, VK;Suthanthiran, M
通讯作者：
Suthanthiran, M
影响因子：
2.7
作者：
Dunmire, V;Wu, CL;Zhang, W
通讯作者：
Zhang, W