Lab-on-a-Film disposable for genotyping multidrug-resistant Mycobacterium tuberculosis from sputum extracts.
Lab-on-a-Film disposable for genotyping multidrug-resistant Mycobacterium tuberculosis from sputum extracts.
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DOI：
10.1039/c8lc01404c
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发表时间：
2019-03
期刊：
影响因子：
6.1
通讯作者：
Alexander C Kukhtin;T. Sebastian;J. Golova;A. Perov;C. Knickerbocker;Yvonne Linger;A. Bueno;Peter Qu;Michael Villanueva;Rebecca C Holmberg;D. Chandler;C. Cooney
中科院分区：
文献类型：
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作者：
Alexander C Kukhtin;T. Sebastian;J. Golova;A. Perov;C. Knickerbocker;Yvonne Linger;A. Bueno;Peter Qu;Michael Villanueva;Rebecca C Holmberg;D. Chandler;C. Cooney
我们描述了一种一次性薄膜实验室，可从痰液提取物中检测耐多药结核病（MDR-TB）。一次性薄膜实验室由203个凝胶元件组成，其中包括5个基因（包括内部对照）中37个突变、缺失或插入元件的DNA序列（探针）。这些凝胶元件印刷在柔性薄膜上，其成本比微阵列玻璃低约500倍。然后将具有印刷的凝胶元件的膜层压到另外的可卷曲材料（膜）上以形成微流体流动池。我们在单个微流体室中组合了多重扩增和杂交步骤，在杂交之前和整个杂交过程中没有缓冲液交换或其他操作。该流动池还包括杂交后清洗步骤，同时保留完全封闭的扩增子系统，从而最大限度地减少样品或扩增子交叉污染的可能性。我们报告了所有MDR-TB标记物的分析灵敏度为32 cfu mL-1，并使用自动TruTip工作站进行提取和Lab-on-a-Film一次性处理用于扩增和检测提取物检测MDR-TB阳性临床标本。
We describe a Lab-on-a-Film disposable that detects multidrug-resistant tuberculosis (MDR-TB) from sputum extracts. The Lab-on-a-Film disposable consists of 203 gel elements that include DNA sequences (probes) for 37 mutations, deletions, or insertion elements across 5 genes (including an internal control). These gel elements are printed on a flexible film, which costs approximately 500 times less than microarray glass. The film with printed gel elements is then laminated to additional rollable materials (films) to form a microfluidic flow cell. We combined multiplex amplification and hybridization steps in a single microfluidic chamber, without buffer exchanges or other manipulations up to and throughout hybridization. This flow cell also incorporates post hybridization wash steps while retaining an entirely closed-amplicon system, thus minimizing the potential for sample or amplicon cross-contamination. We report analytical sensitivity of 32 cfu mL-1 across all MDR-TB markers and detection of MDR-TB positive clinical specimens using an automated TruTip workstation for extraction and the Lab-on-a-Film disposable for amplification and detection of the extracts.