Biosensors-Basel
·
2024
·Vol. 14
·期 2
·105
·
IF 4.9
Xinmeng Zhang,Cheng Zhou,Jianxun Hou,Gang Feng,Zhourui Xu,Yonghong Shao,Chengbin Yang,Gaixia Xu
Precise DNA quantification and nuclear imaging are pivotal for clinical testing, pathological diagnosis, and drug development. The detection and localization of mitochondrial DNA serve as crucial indicators of cellular health. We introduce a novel conjugated oligoelectrolyte (COE) molecule, COE-S3, featuring a planar backbone composed of three benzene rings and terminal side chains. This unique amphiphilic structure endows COE-S3 with exceptional water solubility, a high quantum yield of 0.79, and a significant fluorescence Stokes shift (λex= 366 nm, λem= 476 nm), alongside a specific fluorescence response to DNA. The fluorescence intensity correlates proportionally with DNA concentration. COE-S3 interacts with double-stranded DNA (dsDNA) through an intercalation binding mode, exhibiting a binding constant (K) of 1.32 × 106M−1. Its amphiphilic nature and strong DNA affinity facilitate its localization within mitochondria in living cells and nuclei in apoptotic cells. Remarkably, within 30 min of COE-S3 staining, cell vitality can be discerned through real-time nuclear fluorescence imaging of apoptotic cells. COE-S3’s high DNA selectivity enables quantitative intracellular DNA analysis, providing insights into cell proliferation, differentiation, and growth. Our findings underscore COE-S3, with its strategically designed, shortened planar backbone, as a promising intercalative probe for DNA quantification and nuclear imaging.
研究领域:
生物技术
·
分子生物学
·
医学成像
·
细胞生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Sensors & Diagnostics
·
2024
·Vol. 3
·期 12
·1984-1991
·
IF 3.5
Jinrong Chen,Ruwen Xie,Rui Liu,Lishang Liu,Shusheng Zhang
Visual analysis methods have received widespread attention due to their simplicity, economy, and intuitive results. In this work, a visual DNA quantitative analysis method based on surface selective site-directed crystallization (SSSC) was developed. Firstly, we explored the formation of calcium carbonate crystals with unique polymorphism induced by the surface of functionalized glass slides with different groups; among them, the calcite induced by the –COOH functional group has a uniform shape, larger size, and even distribution, so it serves as a signal promoter. In contrast, due to the –N(CH3)3 group acting as a signal inhibitory molecule by inhibiting crystallization, the signal molecule is captured through DNA hybridization, and the crystallization reaction is performed. The calcite growing on the DNA site is visible to the naked eye, and the DNA molecules hybridized on the surface of the glass slide are further quantified. The detection limit of this proposed visual method is 0.1 fM, and only a smartphone is needed to complete basic quantification. This work provides a basis for research into the use of single crystals as digital readouts in the field of DNA analysis, with the advantages of being simple and economical and requiring minimal equipment.
研究领域:
生物化学
·
分子生物学
·
分析化学
✓ 已验证
产品关联
DOI
高影响因子
LAB ON A CHIP
·
2023
·
IF 6.1
xiaoliang zhang,Shun Wang,Jinxian Wang,Xiaojie Sun,Jinbing Xue,Zhenya Wang,Tianhang Yang,Liangfei Weng,Bidou Wang,Gangyin Luo
Droplet digital PCR (ddPCR) is a powerful method for absolute nucleic acid quantification with high precision and accuracy. However, complicated operational steps have hampered the use and diffusion of ddPCR. Therefore, an automated, easy-to-use, low-sample-consumption, and portable ddPCR platform is urgently needed. This paper proposes a microfluidic ddPCR platform based on a microfluidic chip that can realize the sample-to-result function by switching the rotary valve, achieving the dual function of the flow-focusing structure for droplet generation and readout. Sample, generation oil, and analysis oil were pre-added to the reservoirs. Droplets were generated due to focusing flow, and after passing through the integrated temporary storage bin in the rotary valve, the droplets and oil subsequently entered the collecting tube, improving the droplet-to-oil volume ratio for enhanced thermal cycle performance. Droplets with an average diameter of 107.44 μm and a CV of 2.38% were generated using our chip under the optimal pressures. High-performance thermal cycling was achieved through improvements of the droplet-to-oil volume ratio of the sample, the integrated heating lid, the pure copper heating base, and the temperature-controlling algorithm. Gradient quantification experiments were conducted for the HER2 and CEP17 genes extracted from breast cancer cells, yielding strong linear correlations with R2 values of 0.9996 for FAM and 0.9989 for CY5. Moreover, pronounced linearity was obtained between the detected concentrations of HER2 and CEP17, indicated by a slope of 1.0091 and an R² of 0.9997, signifying consistent HER2:CEP17 ratios across various sample dilutions. The outcomes of the quantitative analysis, encompassing the dynamic range and the consistency of the HER2:CEP17 ratio using our ddPCR platform, meet the standards required for breast cancer assessment and therapy. Our ddPCR platform is automated, portable, and capable of stable droplet generation, high-efficiency amplification, realization of the sample-to-result function based on dual-function flow-focusing structure, and accuracy absolute quantification, underscoring its significant potential for ddPCR analysis in clinical diagnostics.
研究领域:
生物医学工程
·
分子生物学
·
临床诊断
✓ 已验证
产品关联
DOI
PubMed
高影响因子
CHEMOMETRICS AND INTELLIGENT LABORATORY SYSTEMS
·
2015
·
IF 2.321
Jing-Jing Kong,Zeng-Ping Chen,Yao Chen,Xiu-Fang Yan,Ru-Qin Yu
Accurate quantification of genomic DNA is fundamental to many molecular analyses such as diagnostic assays and genotyping. Due to relatively low and varying extraction rates of DNA from cell lysates by genomic DNA extraction kits, it is desired to quantitatively determine DNA contents directly in cell lysates. However, the coexistence of scatterers (e.g., cell debris), absorbers, and intrinsic fluorophores in cell lysates prevents either UV–Vis spectrophotometric methods or fluorometric methods based on fluorescent dyes and ratiometric models to be directly applied to cell lysates. In this contribution, a novel fluorometric method based on an advanced chemometric model which can effectively eliminate the effects of scatterers and absorbers on fluorescence spectra of cell lysates was proposed for the direct quantification of DNA in lysates of CEM cells. Without cumbersome prior DNA extraction, the proposed method achieved quite stable and accurate DNA quantification with recovery rates in the range of 89%–115%, considerably better than that of either the fluorometric method based on the ratiometric model or the UV–Vis spectrophotometric method coupled with DNA extraction. The proposed method has the advantages of rapidness, simplicity, and high precision, and can be developed into a promising alternative for accurate quantification of genomic DNA.
研究领域:
分子生物学
·
生物化学
·
分析化学
✓ 已验证
产品关联
DOI
高影响因子
危险品化学品经营许可证（带存储）