Development of Carbon Dots-Based Nanosomes as Fluorescent Cell Labeling Reagents

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Project title

Code/計畫編號

MOST108-2622-M019-002-CC2

Translated Name/計畫中文名

開發碳量子點奈米微脂囊的細胞螢光標定試劑

Funding Organization/主管機關

National Science and Technology Council

Co-Investigator(s)/共同執行人

張煥宗

Department/Unit

Department of Bioscience and Biotechnology

Website

https://www.grb.gov.tw/search/planDetail?id=13238298

Year

2019

Start date/計畫起

01-11-2019

Expected Completion/計畫迄

31-10-2020

Bugetid/研究經費

436千元

ResearchField/研究領域

化學

本產學計畫為開發新穎碳量子奈米微脂囊 (carbon-dot nanosomes; CDNSs) 於細胞結構、胞器 (organelle)、次胞器 (sub-organelle) 的螢光標定。傳統細胞螢光染劑具有染色步驟耗時、單一激發波長限制、光穩定性不佳以及毒性等問題而限制了其廣用性。目前常見的細胞螢光染劑，如針對細胞核 (nucleus) 染色的Hoechst和DAPI染劑、細胞膜 (plasma membrane) 的DiO系列染劑、肌動蛋白絲 (actin filaments) 之鬼筆環肽 (Phalloidin) 類和核仁 (nucleolus) 之RNA染劑 (SYTO RNASelect Green Fluorescent cell Stain)，然而這些染劑之光穩定性皆不佳。再者，Phalloidin類染劑需繁瑣之細胞固定和破膜步驟才能進行染色，無法應用於活細胞染色。而DiO系列染劑和核仁之RNA染劑，其本身易於聚集，需現配現用或經過費時的加熱再溶解、分散步驟，使用上較為費時且不便。近年，市面上出現利用胜肽修飾之金屬量子點 (quantum dots, QDs) 做為細胞質之螢光染劑，如Qtracker系列染劑，與傳統細胞螢光染劑相比，雖有較佳之光穩定性，但由於金屬量子點常由重金屬、類金屬元素，如: 鎘、鋅、鉛、銦或硒、碲等組成，具細胞毒性，其生物相容性尚有疑慮。為解決市售螢光染劑尚存之問題，我們將透過與森兆生醫合作開發，以綠色化學合成法製備、分離一系列親/疏水性各異的碳量子點納米微脂囊，利用其容易以細胞膜融合進入細胞的特性，達到對細胞不同結構、胞器、次胞器的快速標定。並應用其多顏色螢光、光穩定性佳、生物相容性佳和子代細胞可傳遞性等特點，能在不同的激發光源下實現細胞結構、胞器、次胞器的動態、多代數觀察。 In this industry-university cooperative research project, we aim to develop novel carbon-dot nanosomes (CDNSs) for labeling of cellular structures and sub-organelles. It still remains remains some challenges when using traditional cellular fluorescent stains, including time-consuming procedure, limitation to a single excitation wavelength, poor photostability, and cytotoxicity. Common cellular fluorescent stains, such as Hoechst and DAPI (for nucleus), DiO series stains (for plasma membrane), phalloidin-based stains (for actin filaments), RNA dyes-based nucleolus stains (SYTO RNASelect Green Fluorescent Cell Stain) are prone to photobleaching. Besides, phalloidin-based stains require complicated steps, including cell fixation and membrane permeabilization, for successful staining; and the stains cannot be used to stain live cells. The series of DiO stains and nucleolar RNA stains are tend to aggregation and must use immediately after preparation; otherwise, additional steps, through heating and re-dissolution, are required for better dispersity. Recently, peptide-modified quantum dots (QDs), such as Qtracker stains, have been utilized as cytoplasmic stains, which have better photostability when compared to traditional cellular stains. Howevr, metallic quantum dots are usually consist of heavy metal and metalloid elements, such as cadmium, zinc, lead, indium or selenium, tellurium, etc., which are cytotoxic and arising concerns over biocompatibility. To solve these problems, we are in cooperation with Bio-helix Co., Ltd. to synthesize and prepare a series of CDNSs with varying hydrophilicity/hydrophobicity by green chemistry approach. Taking advantages of the lipophilic properties, CDNSs can achieve rapid and high efficient staining of cellular structures and sub-organells through membrane fusion. The CDNSs, with excellent photostability, well biocompatibility, and generational transferability, can realize multi-color dynamic and multi-generational imaging of cellular structures and sub-organelles under multiple excitation wavelengths.

Keyword(s)

碳量子點納米微脂囊
次胞器
細胞標定
多顏色螢光
光穩定性
carbon-dot nanosomes
sub-organelles
cell labeling
multi-color fluorescence
photostability

DSpace CRIS

Development of Carbon Dots-Based Nanosomes as Fluorescent Cell Labeling Reagents

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