The separation of exfoliated cells from the brushes used during cervico-vaginal smears is difficult, a problem which may affect the quality of ribonucleic acid (RNA) extracted. We compared the results of RNA extraction from cervico-vaginal cytology samples according to the type of tubes, preservative solutions, and storage temperature. The samples included exfoliated cervico-vaginal cytological specimens from patients with human papilloma virus 16, positive for cervical intraepithelial neoplasia or cervical cancer. Exfoliated cells were obtained by shaking a brush in a conventional rigid vial tube or squeezing the brush in a soft vial tube. RNA quantity and quality were compared between the two tubes. The concentration and purity of RNA (A260/A280 and A260/A230 ratios) was compared amongst five groups: Group 1, standard frozen storage; Group 2-4, RNA stabilization reagents with room temperature [RNAlater RNA Stabilization Reagent, RNAprotect cell Reagent and AllProtect Tissue Reagent]; and Group 5, Surepath Preservative fluid. To demonstrate the utility of the extracted RNA for PCR-based cDNA synthesis, GAPDH and E6 were targeted and gel band densities of GAPDH and E6 were measured. The median RNA concentration was significantly higher in the soft tubes compared with the rigid tubes (100.2 vs. 7.1 ng/μL, p = 0.0209). The purity of the RNA was higher in soft vial tubes than in rigid vials, as measured by A260/280 and A260/230 ratios. The RNA concentration, purity, and GAPDH density of groups 1, 2 and 3 were significantly higher than those of groups 4 and 5. Moreover, E6 density of group 1 and 2 was significantly higher than that of group 3, 4 and 5. The use of soft tubes enhanced the mRNA quantity and quality in cervico-vaginal cytology. The products of mRNA extraction using RNAlater RNA Stabilization Reagent and RNAprotect Cell Reagent at room temperature were comparable to those obtained by conventional frozen storage. Our protocol improved the yield and quality of RNA and might produce better results for molecular analysis in cervico-vaginal cytology.

中文翻译：

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宫颈阴道细胞学中RNA数量和质量的提高。

在宫颈阴道涂片检查中很难将脱落的细胞与刷子分开，这个问题可能会影响提取的核糖核酸（RNA）的质量。我们根据管的类型，防腐剂溶液和保存温度比较了宫颈细胞学样本中RNA提取的结果。样本包括来自人乳头瘤病毒16型患者的宫颈宫颈脱落细胞学标本，对宫颈上皮内瘤变或宫颈癌呈阳性。通过在常规的刚性小瓶管中摇动刷子或在软的小瓶管中挤压刷子来获得脱落的细胞。比较了两个试管之间的RNA数量和质量。在五个组中比较了RNA的浓度和纯度（A260 / A280和A260 / A230的比例）：第1组，标准冷冻保存；2-4组 具有室温的RNA稳定剂[RNAlater RNA稳定剂，RNAprotect细胞试剂和AllProtect组织试剂]；第5组，Surepath防腐液。为了证明提取的RNA在基于PCR的cDNA合成中的实用性，我们以GAPDH和E6为目标，并测量了GAPDH和E6的凝胶条带密度。与刚性试管相比，软试管中的RNA浓度中位数显着更高（100.2对7.1 ng /μL，p = 0.0209）。通过A260 / 280和A260 / 230的比值测量，在软小瓶试管中，RNA的纯度高于刚性小瓶。第1、2和3组的RNA浓度，纯度和GAPDH密度显着高于第4和5组。此外，第1和2组的E6密度显着高于第3、4和5组。软管的使用增强了子宫颈阴道细胞学中的mRNA数量和质量。在室温下使用RNAlater RNA稳定试剂和RNAprotect Cell试剂提取mRNA的产物与常规冷冻保存的产物相当。我们的协议提高了RNA的产量和质量，并可能在宫颈阴道细胞学分子分析中产生更好的结果。