School of Nano-Bioscience and Chemical Engineering, Ulsan National Institute of Science and Technology, Banyeon-ri 100, Ulsan 689-798, Republic of Korea.
Biosens Bioelectron. 2013 Aug 15;46:175-82. doi: 10.1016/j.bios.2013.02.038. Epub 2013 Mar 4.
Bacterial bioreporters are limited in their abilities to detect large polar molecules due to their membrane selectivity. In this study, the activity of serum complement was used to bypass this undesired selectivity. Initially, the serum complement activity was assessed using the responses of a bacterial bioreporter harboring a recA::luxCDABE transcriptional fusion when exposed to the chemotherapy drug, mitomycin C (MMC). Using 50 °C-treated serum, the limit of detection for this bacterial sensor was lowered by nearly 450-fold, from 31 μg/L to 0.07 μg/L MMC. Real-time quantitative PCR demonstrated that serum-treated cultures responded more strongly to 100 μg/L MMC, with 3.1-fold higher recA expression levels. Subsequent experiments with other bioreporter strains also found enhanced sensitivities and responses. Finally, combining each of the above findings, tests were performed to demonstrate the potential application of the recA::luxCDABE bioreporter within a lab-on-a-CD platform as a point-of-care diagnostic to measure chemotherapeutic drug concentrations within blood.
由于细胞膜的选择性，细菌生物报告器在检测大极性分子方面的能力受到限制。在这项研究中，血清补体的活性被用来绕过这种不理想的选择性。最初，使用携带 recA::luxCDABE 转录融合的细菌生物报告器对血清补体活性进行了评估，当该生物报告器暴露于化疗药物丝裂霉素 C（MMC）时，会产生响应。使用 50°C 处理的血清，该细菌传感器的检测限降低了近 450 倍，从 31μg/L 降至 0.07μg/L MMC。实时定量 PCR 表明，经血清处理的培养物对 100μg/L MMC 的反应更强烈，recA 表达水平高出 3.1 倍。随后对其他生物报告器菌株进行的实验也发现了灵敏度和响应的提高。最后，结合上述发现，进行了测试以证明 recA::luxCDABE 生物报告器在 CD 片上实验室平台中的潜在应用，作为一种即时诊断方法，用于测量血液中的化疗药物浓度。
