Label-free recombinase polymerase amplification with hyperspectral digital optofluidics DOI Open Access
Tianqi Zhou, Xiangyu Jin, F. Yang

и другие.

Опубликована: Окт. 12, 2023

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

Язык: Английский

A review of electrochemical sensing in droplet systems: droplet and digital microfluidics DOI Creative Commons
Kieu Chan Ly, Abrarkhan M. Pathan, Darius G. Rackus

и другие.

Analytica Chimica Acta, Год журнала: 2025, Номер 1347, С. 343744 - 343744

Опубликована: Фев. 4, 2025

Язык: Английский

Процитировано

1
Droplet-Based Microfluidics with Mass Spectrometry for Microproteomics DOI Creative Commons
Hang Li,

Yudan Ma,

Rongxin Fu

и другие.

Engineering, Год журнала: 2024, Номер unknown

Опубликована: Сен. 1, 2024

Язык: Английский

Процитировано

4
Simple In-Cell Processing Enables Deep Proteome Analysis of Low-Input Caenorhabditis elegans DOI Creative Commons

Malek Elsayyid,

Jessica E. Tanis, Yanbao Yu

и другие.

Analytical Chemistry, Год журнала: 2025, Номер unknown

Опубликована: Апрель 21, 2025

Caenorhabditis elegans is a widely used genetic model organism; however, the worm cuticle complicates extraction of intracellular proteins, prerequisite for typical bottom-up proteomics. Conventional physical disruption procedures are not only time-consuming but can also cause significant sample loss, making it difficult to perform proteomics with low-input samples. Here, first time, we present an on-filter in-cell (OFIC) processing approach that digest C. proteins directly in cells organism after methanol fixation. With OFIC and single-shot LC-MS analysis, identified over 9400 from 200 worms, largest proteome reported date did require fractionation or enrichment. We systematically evaluated performance by comparing conventional lysis-based methods. Our data suggest superior identification quantitation. further even lower-input samples, including single worms. Then, this method determine how impacted loss superoxide dismutase sod-1, ortholog human SOD1, gene associated amyotrophic lateral sclerosis. Analysis 8800 50 worms as initial input showed sod-1 affects abundance required stress response, ribosome biogenesis, metabolism. In conclusion, our streamlined approach, which be broadly applied other systems, minimizes while offering simplest workflow

Язык: Английский

Процитировано

0
Digital Microfluidics for Sample Preparation in Low‐Input Proteomics DOI Creative Commons

Max K. Steinbach,

Jan Leipert,

Theo Matzanke

и другие.

Small Methods, Год журнала: 2024, Номер unknown

Опубликована: Авг. 29, 2024

Abstract Low‐input proteomics, also referred to as micro‐ or nanoproteomics, has become increasingly popular it allows one elucidate molecular processes in rare biological materials. A major prerequisite for the analytics of minute protein amounts, e.g., derived from low cell numbers, down single cells, is availability efficient sample preparation methods. Digital microfluidics (DMF), a technology allowing handling and manipulation liquid volumes, recently been shown be powerful versatile tool address challenges low‐input proteomics. Here, an overview provided on recent advances proteomics using DMF. In particular, capability DMF isolate proteomes cells small model organisms, perform all necessary chemical steps, such denaturation proteolytic digestion on‐chip, are highlighted. Additionally, prerequisites making these steps compatible with follow‐up analytical methods chromatography‐mass spectrometry will discussed.

Язык: Английский

Процитировано

1
Microproteomic-Based Analysis of the Goat Milk Protein Synthesis Network and Casein Production Evaluation DOI Creative Commons
Li Chen, Hiroaki Taniguchi, Emilia Bagnicka

и другие.

Foods, Год журнала: 2024, Номер 13(4), С. 619 - 619

Опубликована: Фев. 19, 2024

Goat milk has been consumed by humans since ancient times and is highly nutritious. Its quality mainly determined its casein content. Milk protein synthesis controlled a complex network with many signal pathways. Therefore, the aim of our study to clearly depict pathways involved in goat mammary epithelial cells (GMECs) using state-of-the-art microproteomic techniques identify key genes pathway. The analysis identified more than 2253 proteins, 323 annotated from proteins. Knockdown IRS1 expression significantly influenced composition (α, β, κ); therefore, this also examined insulin receptor substrate 1 (IRS1) gene closely. A total 12 differential proteins (DEPs) were characterized as upregulated or downregulated IRS1-silenced sample compared negative control. enrichment these DEPs GMECs GO annotation KEGG, well KOG analysis. Our findings expand understanding functional goats, paving way for new approaches modifying content dairy industry product development.

Язык: Английский

Процитировано

0
In-cell processing enables rapid and in-depth proteome analysis of low-input Caenorhabditis elegans DOI Creative Commons

Malek Elsayyid,

Jessica E. Tanis, Yanbao Yu

и другие.

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Сен. 19, 2024

Abstract Caenorhabditis elegans is a widely used genetic model organism, however, the worm cuticle complicates extraction of intracellular proteins, prerequisite for typical bottom-up proteomics. Conventional physical disruption procedures are not only time-consuming, but can also cause significant sample loss, making it difficult to perform proteomics with low-input samples. Here, first time, we present an on-filter in-cell (OFIC) processing approach, which digest C. proteins directly in cells organism after methanol fixation. With OFIC and single-shot LCMS analysis, identified over 9,400 from 200 worms, largest proteome reported date that did require fractionation or enrichment. We systematically evaluated performance approach by comparing conventional lysis-based methods. Our data suggest equivalent unbiased identification quantitation. further even lower input samples, then this method determine how impacted loss superoxide dismutase sod-1 , ortholog human SOD-1 gene associated amyotrophic lateral sclerosis (ALS). Analysis 8,800 50 worms as initial showed affects abundance required stress response, ribosome biogenesis, metabolism. In conclusion, our streamlined be broadly applied other systems, minimizes while offering simplest workflow analysis.

Язык: Английский

Процитировано

0
Label-free recombinase polymerase amplification with hyperspectral digital optofluidics DOI Open Access
Tianqi Zhou, Xiangyu Jin, F. Yang

и другие.

Опубликована: Окт. 12, 2023

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

Язык: Английский

Процитировано

0