Single cell capture, isolation, and long‐term in‐situ imaging using quantitative self‐interference spectroscopy

Fu, Rongxin; Su, Ya; Wang, Ruliang; Xue, Lan; Jin, Xiangyu; Yang, Han Na; Du, Wenli; Shan, Xiuming; Lv, Wenqi; Huang, Guanwen

doi:10.1002/cyto.a.24333

Cited by 12 publications

(9 citation statements)

References 41 publications

(36 reference statements)

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“…The interferometric spectra and statistical results are shown in Figure C,D. To determine the limit of detection (LOD), three times the standard deviation divided by the fitting slope was calculated . Therefore, the LOD for pure dsDNA samples was 3 × 0.10/0.0895 = 3.35 ng/μL.…”

Section: Resultsmentioning

confidence: 99%

Microfluidic Biosensor for Rapid Nucleic Acid Quantitation Based on Hyperspectral Interferometric Amplicon-Complex Analysis

Jin

et al. 2021

ACS Sens.

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Nucleic acid detection plays a vital role in both biomedical research and clinical medicine. The temperature circulation changes of the widely used polymerase chain reaction technique are time-consuming and technically challenging for system development. Recombinase polymerase amplification (RPA) is an isothermal method for rapid nucleic acid detection. However, current RPA amplicon detection methods are complicated and expensive and easily generate false positives, restricting the promotion of RPA techniques. In this work, a hyperspectral interferometric amplicon-complex quantitation method is presented, combined with asymmetric dipole complex strategy optical scattering analysis. GelRed dye was utilized to form amplicon-complex particles, and the Fourier domain spectrum computation contributed to complex scattering quantitation. With this method, a supporting microfluidic chip and automatic system were developed to achieve integrated, rapid, quantitative, and miniscule nucleic acid detection. The Plasmodium falciparum dhfr gene was utilized as an example for targeted nucleic acid quantitation and single nucleotide polymorphism detection. The total reaction time was decreased to merely 20 min, and the limit of detection was only 3.17 ng/μL. The minimum measurable concentration of target was 1.68 copies/ μL, 31.67 times more sensitive than turbidity detection, and the single reaction chamber was only 9.33 μL. No scattering increase occurred for template-free control, and thus, false positives caused by primer dimers and nonspecific products could be avoided. The experimental results prove that the provided method and system can detect single-base mutations in the dhf r gene and is a reasonable technique for rapid, automatic, and low-cost nucleic acid detection.

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Section: Resultsmentioning

confidence: 99%

Microfluidic Biosensor for Rapid Nucleic Acid Quantitation Based on Hyperspectral Interferometric Amplicon-Complex Analysis

Jin

et al. 2021

ACS Sens.

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“…In addition, dry cell mass remained nearly constant during osmotic pressure variations, whereas cellular size changed dramatically. This means that MWII could reveal the intrinsic cellular status without culture environment fluctuation disturbance, which is a potential merit for biological applications [ 30 ].…”

Section: Discussionmentioning

confidence: 99%

Label-Free and Quantitative Dry Mass Monitoring for Single Cells during In Situ Culture

et al. 2021

Cells

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Quantitative measurement of single cells can provide in-depth information about cell morphology and metabolism. However, current live-cell imaging techniques have a lack of quantitative detection ability. Herein, we proposed a label-free and quantitative multichannel wide-field interferometric imaging (MWII) technique with femtogram dry mass sensitivity to monitor single-cell metabolism long-term in situ culture. We demonstrated that MWII could reveal the intrinsic status of cells despite fluctuating culture conditions with 3.48 nm optical path difference sensitivity, 0.97 fg dry mass sensitivity and 2.4% average maximum relative change (maximum change/average) in dry mass. Utilizing the MWII system, different intrinsic cell growth characteristics of dry mass between HeLa cells and Human Cervical Epithelial Cells (HCerEpiC) were studied. The dry mass of HeLa cells consistently increased before the M phase, whereas that of HCerEpiC increased and then decreased. The maximum growth rate of HeLa cells was 11.7% higher than that of HCerEpiC. Furthermore, HeLa cells were treated with Gemcitabine to reveal the relationship between single-cell heterogeneity and chemotherapeutic efficacy. The results show that cells with higher nuclear dry mass and nuclear density standard deviations were more likely to survive the chemotherapy. In conclusion, MWII was presented as a technique for single-cell dry mass quantitative measurement, which had significant potential applications for cell growth dynamics research, cell subtype analysis, cell health characterization, medication guidance and adjuvant drug development.

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“…Here, ∆ obtained contains thickness information of the double-stranded DNA layer, and its corresponding calculation formula is ∆ = 2𝑛𝐷𝑁𝐴𝜎, defined optically as the optical path difference [16] . Considering that in practical situations, the double-stranded DNA layer formed by the amplification product is not dense, and the average refractive index of the double-stranded DNA layer is not easily measured directly, we select the optical path difference ∆ as the physical quantity representing the double-stranded DNA layer.…”

Section: Label-free Detection With Hyperspectral Interferencementioning

confidence: 99%

Label-free recombinase polymerase amplification with hyperspectral digital optofluidics

Zhou,

Jin,

Yang

et al. 2023

Optics in Health Care and Biomedical Optics XIII

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Nucleic acid detection techniques have played a crucial role in identifying specific genetic indicators or species, with Polymerase Chain Reaction (PCR) being the established gold standard in this field. However, PCR's dependence on specialized equipment and skilled personnel has limited its utility in resource-limited or field settings, and its multitemperature stage protocol hinders rapid nucleic acid detection. The emergence of isothermal amplification methods, particularly recombinase polymerase amplification (RPA), has addressed some of these limitations, offering high sensitivity and efficiency. Nevertheless, the challenge of RPA amplicon detection, typically reliant on labeling methods, has persisted, potentially introducing false positives and increased costs. This study introduces an innovative approach to nucleic acid detection, harnessing hyperspectral quantitative interference for label-free, isothermal nucleic acid detection within a remarkably short 25-minute timeframe. By employing a solid-phase RPA amplification process that transforms the product into a DNA molecule layer, and leveraging Fourier domain optical slice separation and spectral phase shift analysis, this method enables the semi-quantitative determination of amplification results. The integration of digital microfluidic technology further enhances the method's performance, enabling parallel, integrated, and clinical multiindicator pathogen detection. Overall, this research presents a practical and rapid solution for label-free nucleic acid detection, addressing the current limitations associated with nucleic acid amplification techniques. This advancement holds promise for a wide range of applications, from point-of-care diagnostics to field-based pathogen detection, ultimately contributing to more accessible and efficient nucleic acid testing methodologies.

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Single cell capture, isolation, and long‐term in‐situ imaging using quantitative self‐interference spectroscopy

Cited by 12 publications

References 41 publications

Microfluidic Biosensor for Rapid Nucleic Acid Quantitation Based on Hyperspectral Interferometric Amplicon-Complex Analysis

Microfluidic Biosensor for Rapid Nucleic Acid Quantitation Based on Hyperspectral Interferometric Amplicon-Complex Analysis

Label-Free and Quantitative Dry Mass Monitoring for Single Cells during In Situ Culture

Label-free recombinase polymerase amplification with hyperspectral digital optofluidics

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