Quantitative analysis of transferrin uptake into living cells at single-molecule level.

IF 5.6 1区化学 Q1 CHEMISTRY, ANALYTICAL Talanta Pub Date : 2025-01-01 Epub Date: 2024-10-18 DOI:10.1016/j.talanta.2024.127031

Marta Pilz, Tomasz Kalwarczyk, Krzysztof Burdzy, Robert Hołyst

引用次数: 0

Abstract

Endocytosis, apart from providing a mechanism for cells to take up nutrients, is the principal route of entry for many nanomedicines. The evaluation of therapeutic delivery efficiency without providing quantitative parameters, like the number of molecules entering the cell and the time of their entry, is very limited. Despite advances in single-molecule methods for in-cell quantitative measurements, they have not become widely used in the study of endocytosis. Their application, however, is challenging owing to the appropriate experimental design and applicable analysis methods. Here, by using an integrated approach based on the multidimensional time-correlated single-photon counting (TCSPC) technique, we demonstrate the quantitative method to measure the time- and concentration-dependent uptake of TRITC-transferrin into living cells with single-molecule sensitivity. The methodology opens possibilities for quantitatively assessing the delivery efficiency of fluorescent molecules into living cells.

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在单分子水平上定量分析活细胞对转铁蛋白的吸收。

内吞作用除了为细胞提供摄取营养物质的机制外，还是许多纳米药物进入细胞的主要途径。如果不提供定量参数，如进入细胞的分子数量和进入细胞的时间，对治疗递送效率的评估就非常有限。尽管用于细胞内定量测量的单分子方法取得了进展，但这些方法尚未广泛应用于内吞研究。然而，由于适当的实验设计和适用的分析方法，这些方法的应用具有挑战性。在这里，我们利用基于多维时间相关单光子计数（TCSPC）技术的综合方法，展示了一种定量方法，以单分子灵敏度测量活细胞对TRITC-铁蛋白随时间和浓度变化的摄取。该方法为定量评估荧光分子进入活细胞的效率提供了可能。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Talanta 化学-分析化学

CiteScore

12.30

自引率

4.90%

发文量

861

审稿时长

29 days

期刊介绍： Talanta provides a forum for the publication of original research papers, short communications, and critical reviews in all branches of pure and applied analytical chemistry. Papers are evaluated based on established guidelines, including the fundamental nature of the study, scientific novelty, substantial improvement or advantage over existing technology or methods, and demonstrated analytical applicability. Original research papers on fundamental studies, and on novel sensor and instrumentation developments, are encouraged. Novel or improved applications in areas such as clinical and biological chemistry, environmental analysis, geochemistry, materials science and engineering, and analytical platforms for omics development are welcome. Analytical performance of methods should be determined, including interference and matrix effects, and methods should be validated by comparison with a standard method, or analysis of a certified reference material. Simple spiking recoveries may not be sufficient. The developed method should especially comprise information on selectivity, sensitivity, detection limits, accuracy, and reliability. However, applying official validation or robustness studies to a routine method or technique does not necessarily constitute novelty. Proper statistical treatment of the data should be provided. Relevant literature should be cited, including related publications by the authors, and authors should discuss how their proposed methodology compares with previously reported methods.