期刊
ANALYTICAL CHEMISTRY
卷 79, 期 16, 页码 6135-6142出版社
AMER CHEMICAL SOC
DOI: 10.1021/ac0703698
关键词
-
资金
- NHGRI NIH HHS [R01 HG002613] Funding Source: Medline
In this report, we show that a novel capillary-based photopolymerized monolith offering unprecedented efficiency (similar to 80%) for DNA extraction from submicroliter volumes of whole blood (Wen, J.; Guillo, C.; Ferrance, J. P.; Landers, J. P. Anal. Chem. 2006, 78, 1673-1681) can be translated to microfluidic devices. However, owing to the large mass of protein present in blood, both DNA binding capacity and extraction efficiency were significantly decreased when extraction of DNA was carried out directly from whole blood (38 +/- 1%). To circumvent this, a novel two-stage microdevice was developed, consisting in a C 18 reversed-phase column for protein capture (stage 1) in series with a monolithic column for DNA extraction (stage 2). The two-stage, dual-phase design improves the capability of the monolith for whole blood DNA extraction by similar to 100-fold. From a 10-mu L load of whole blood containing 350 ng of DNA, 99% (340 +/- 10 ng) traverses the C18 phase while similar to 70% (1020 +/- 45 ug) of protein is retained. A total of 240 +/- 2 ng of DNA was eluted from the secondstage monolith, resulting in an overall extraction efficiency of 69 1%. This provided not only an improvement in extraction efficiency over other chip-based DNA extraction solid phases but also the highest extraction efficiency reported to-date for such sample volumes in a microfluidic device. As an added bonus, the two-stage, dual-phase microdevice allowed the 2-propanol wash step, typically required to remove proteins from the DNA extraction phase for successful PCR, to be completely eliminated, thus streamlining the process without affecting the PCR amplifiability of the extracted DNA.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据