PAXgene Blood RNA Kit
从稳定于PAXgene Blood RNA Tubes的全血中分离和纯化胞间RNA
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PAXgene Blood RNA Kit (50)
Cat. No. / ID: 762174
50 PAXgene Spin Columns, 50 PAXgene Shredder Spin Columns, Processing Tubes, RNase-Free DNase I, RNase-Free Reagents and Buffers. To be used in conjunction with PAXgene Blood RNA Tubes
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On May 26, 2022, the five-year transitional period (announced in April 2017) for the new EU Regulation on In Vitro Diagnostic Medical Devices (IVDR 2017/746) will end. This requires a transition from EU IVD Directive 98/79/EC (IVDD) to IVDR for all Class A non-sterile products including nucleic acid isolation kits. The completion will be documented by the signing of a Declaration of Conformity (DoC). Once the DoC has been signed, it may take several months for the IVDR product to be commercially available due to international regulatory product registrations outside of the European Union. For those countries, PAXgene Blood RNA products previously CE-marked according to IVDD will be available after May 26, 2022, until the sell-off provision* is reached that is 26th May 2025.
*Only able to sell the IVDD stock that are in the distribution center or warehouse.
These products are not available in all countries; please inquire.
PAXgene Blood RNA Kit 旨在用于体外诊断用途。
✓ 全天候自动处理在线订单
✓ 博学专业的产品和技术支持
✓ 快速可靠的（再）订购
特点
- 符合欧洲体外诊断98/79/EC指令
- 与采样、稳定及纯化形成整合的系统
- 在18–25°C条件下可稳定多至3天
- 在–20°C或–70°C条件下至少稳定60个月
- 分析前标准化的样本处理过程
产品详情
PAXgene Blood RNA系统包括用于血液采集、稳定和运输的PAXgene Blood RNA Tubes（由BD提供，货号762165），以及基于硅胶膜技术、离心柱法分离纯化的PAXgene Blood RNA Kit。纯化可使用离心机进行手工操作，也可在QIAcube全自动核酸纯化仪上自动运行。该产品表现卓越，确保高度可靠的RNA纯化。
绩效
PAXgene Blood RNA Tubes适用于采集全血并稳定细胞内RNA，在18–25°C条件下可稳定长达3天（参见"RNA stability at 18–25°C: FOS"和"RNA stability at 18–25°C: IL1B"），或在2–8°C条件下稳定长达5天（参见"RNA stability at 2–8°C: FOS"和"RNA stability at 2–8°C: IL1B"）。当前的数据显示，细胞内RNA在–20°C或–70°C条件下可保存至少60个月。
使用PAXgene Blood RNA System纯化的总RNA纯度高，A260/A280 值在1.8和2.2之间，且基因组DNA含量不超过1.0% (w/w)。当使用30%的洗脱液时，至少95%的样品在RT-PCR中没有出现反应抑制的情况。
样品制备的平均时间为90分钟（根据12个样品的制备时间），手工操作只需40分钟。在处理95%以上样品时，2.5 ml健康人类全血的RNA产量不低于3 µg。由于产量高度依赖于供体，单次产量可能会有变动。对于单个供体，PAXgene Blood RNA System 确保产量可重复性高（参见 Reproducible and repeatable RNA purification"和" Repeatability and reproducibility of RNA yield"）以及RT-PCR的可重复性高（参见"Reproducibility between users"、"Reproducibility between kit lots", 和表格），高度适合于临床诊断检测。
对于超过95%的样本，从2.5 ml健康人类全血中获得的RNA产量不少于3 μg。图片"RNA yield and purity — automated processing"显示的是3名实验员使用3批试剂盒自动化处理288个样本的RNA产量。由于使用的是混合血液样本而非单个的PAXgene Blood RNA Tubes，实验结果不能反映单一个体血液样本的RNA预期产量。由于产量高度依赖于供体，单个产量会有差别。
当使用多至30%的洗脱液时，至少95%的样品不会出现RT-PCR抑制。通过对RNA阴性样品（水）中beta-球蛋白和FOS转录子序列与RNA阳性样本（人类全血）进行定量RT-PCR分析，未发现使用自动化流程处理样品中有交叉污染。
使用PAXgene Blood RNA System的自动流程制备的RNA纯度高，没有RT-PCR抑制（见上文），A260/A280值在1.8 和2.2之间。经real-time PCR分析beta-肌动蛋白的序列发现，超过95%的样品的基因组DNA含量不高于1% (w/w) 。图片"RNA yield and purity — automated processing"显示了3个实验员使用3批试剂盒处理288个样品的A260/A280值和相对基因组DNA含量。
如图"Reproducibility between automated and manual protocols"所示，使用PAXgene Blood RNA System自动纯化RNA的流程提供高度可重复的RT-PCR结果，适用于临床诊断检测。
| 检测系统 | FOS/18S rRNA assay | IL1B/18S rRNA assay | ||
|---|---|---|---|---|
| 数据对比 | 平均值 | ± SD | 平均值 | ± SD |
| (ΔΔCT) | (ΔΔCT) | (ΔΔCT) | (ΔΔCT) | |
| 同一使用者使用不同批试剂盒的可重复性 | ||||
| 所有使用者，第一批–第一批 | 0.00 | 0.00 | 0.00 | 0.00 |
| 所有使用者，第一批–第二批 | –0.03 | 0.48 | –0.07 | 0.66 |
| 所有使用者，第一批–第三批 | –0.21 | 0.52 | 0.11 | 0.71 |
| 不同使用者使用同一批试剂盒的可重复性 | ||||
| 所有批次，使用者A–使用者A | 0.00 | 0.00 | 0.00 | 0.00 |
| 所有批次，使用者A–使用者B | –0.46 | 0.44 | –0.06 | 0.69 |
| 所有批次，使用者A–使用者C | –0.31 | 0.60 | –0.15 | 0.71 |
查看图表
原理
PAXgene Blood RNA Kit用于从在PAXgene Blood RNA Tube中采集的2.5 ml人类全血中纯化总RNA，操作简单，可手工操作，也可自动化运行（参见"Manual PAXgene Blood RNA procedure"和" Automated PAXgene Blood RNA procedure"）。
PAXgene Blood RNA Tubes 含有专利申请中的特定试剂，基于RNA固定的专利技术（US Patents 6,602,718 和6,617,170）。这些试剂成分可保护RNA分子，避免其被RNA酶降解，并将基因表达体内变化的改变降至最小。PAXgene Blood RNA Tubes适用于采集全血并稳定细胞内RNA，在18–25°C条件下可稳定长达3天（参见"RNA stability at 18–25°C: FOS"和"RNA stability at 18–25°C: IL1B"），或在2–8°C条件下稳定长达5天（参见"RNA stability at 2–8°C: FOS"和"RNA stability at 2–8°C: IL1B"）。当前的数据显示，细胞内RNA在–20°C或–70°C条件下可保存至少60个月。
查看图表
程序
PAXgene Blood RNA纯化操作流程简单，可手动或自动进行（参见"Manual PAXgene Blood RNA procedure"和" Automated PAXgene Blood RNA procedure"）。
PAXgene Blood RNA纯化手动流程
将重悬的沉淀和蛋白酶K一起溶解在经优化的缓冲液中孵育，进行蛋白质酵解。然后应用PAXgene Shredder离心柱再次离心，匀浆细胞裂解液，并去除剩余的细胞残骸。将流过部分的上清液转移到干净的微型离心管中，加入乙醇调节结合条件，将裂解液装入一个PAXgene RNA离心柱中。快速离心后，RNA会选择性地结合到PAXgene硅胶膜上，杂质则流过。剩余的杂质通过几次高效洗涤去除。在第一次和第二次洗涤步骤之间，使用DNA酶处理膜，以去除微量残余的DNA。洗涤之后，使用洗脱缓冲液洗脱RNA并进行热变性。
PAXgene Blood RNA纯化自动流程
使用QIAcube全自动核酸纯化仪制备样品的流程与手动法相同，您可继续使用PAXgene Blood RNA Kit纯化高品质RNA。
自动化的RNA纯化流程由两个部分组成，"PAXgene Blood RNA Part A"和"PAXgene Blood RNA Part B"，在两个部分之间需要短暂的手工操作。
将经离心、洗涤、重悬的核酸沉淀从PAXgene Blood RNA Tube转移到离心管中，将离心放到QIAcube全自动核酸纯化仪工作台的恒温震荡器中。实验员选择并启动菜单中的"PAXgene Blood RNA Part A"。QIAcube全自动核酸纯化仪会自动运行，直到将RNA洗脱至洗脱缓冲液中。实验员将含有已纯化RNA的离心管转移到QIAcube全自动核酸纯化仪的恒温震荡器中，选择并启动菜单中的"PAXgene Blood RNA Part B"，QIAcube全自动核酸纯化仪会执行热变性。
样本平均制备时间是125分钟（基于制备12个样本的时间），手工操作时间只需约20分钟。
查看图表
应用
该试剂盒与PAXgene Blood RNA Tubes共同使用，可从全血中分离纯化细胞内RNA进行RT‑PCR。
辅助数据和图表
Repeatability and reproducibility of RNA yield.
Blood samples from 30 apparently healthy consented donors were collected in PAXgene Blood RNA Tubes (12 tubes per donor, 360 tubes in total). The contents of the tubes from 3 apparently healthy donors were pooled and subsequently realiquoted into 36 samples. These 36 samples per 3-donor-pool were manually processed by 3 different operators. Each operator used 3 different PAXgene Blood RNA Kit lots for the extraction and processed quadruplicate samples from each of the 10 donor pools. [A] RNA yield and standard deviation for every operator–lot combination. Quadruplicate blood samples from 10 donor pools were processed by 3 different operators (A, B, C) with each of 3 kit lots (1, 2, 3). The mean yields (columns) and standard deviations (error bars) per quadruplicate sample from the same donor pool for different operator and different kit lot are presented. [B] CV of RNA yield per donor pool for all operator–lot combinations (A, B, C; 1, 2, 3) as calculated from the mean yield and standard deviation of the yield shown in part A.
Specifications
| Features | Specifications |
|---|---|
| Format | Spin column |
| Technology | Silica technology |
| Sample amount | 2.5 ml |
| Elution volume | 80 µl (2 x 40 µl) |
| Main sample type | Whole blood |
| Time per run | Manual: 90 min/12 samples; Automated: 151 min/12 samples |
| Yield | ≥3 µg/2.5 ml sample (≥95% of all samples processed). Apparently healthy consented donors with white blood cell counts in range of 4.8 x 10^6 – 1.1 x 10^7 leukocytes/ml. |
| Purification of total RNA, miRNA, poly A+ mRNA, DNA or protein | 1.18-2.2 (A260/A280) RNA; ≤1 % (w/w) genomic DNA (≥95% of all samples processed) |
| Processing | Manual: centrifugation; Automated: QIAcube Connect MDx |
资源
产品介绍与指南 (2)
学术海报 (4)
安全数据表 (1)
实验方案软件 (1)
应用说明 (2)
试剂盒操作手册 (1)
基因表达分析 (1)
Safety Data Sheets (1)
Certificates of Analysis (1)
FAQ
Where can I find additional information for PreAnalytiX PAXgene products?
Can the PAXgene Blood RNA System be used with animal blood samples?
Can the PAXgene Blood RNA Tubes undergo freeze/thaw cycles?
What RNA yields are expected with the PAXgene Blood RNA kit?
Why is a blood collection set required to be used with the PAXgene Blood RNA tube?
What is the integrity of RNA from PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue?
How long can the PAXgene Blood RNA tubes be kept frozen at –20°C or –70°C?
What are possible reasons for blood draws with lower than expected blood volume?
Is there any chance of the PAXgene Blood RNA tube reagent going back into the patient's arm?