Thiazole Orange (solution) 

Thiazole Orange (solution) is an asymmetric anthocyanin dye that can be coupled with oligonucleotides (ONs) to prepare fluorescent hybridization probes. Thiazole Orange has been widely used in biomolecular detection and staining of DNA/ RNA in gels and can be used for reticulocyte analysis. Thiazole orange generates a significant fluorescence enhancement and high quantum yield when it binds with nucleic acids, especially RNA. Thiazole orange can permeate living cell membranes. Thiazole orange can use UV light for detection, but can also be detected with blue light. The excitation and emission of Thiazole orange are λ_ex = 510 nm (488 nm and 470 nm also show strong excitation) and λ_em = 527 nm, respectively^{[1][2][3][4][5][6]}.
Solvent and concentration: DMSO: 10 mM
The 1 mL volume is defined as the base specification. All larger sizes correspond to incremental volumes of this base.

Thiazole Orange 染色液的配制：
1.1 工作液的配制
将 Thiazole Orange 储存液在含有 0.002 M EDTA 和 0.02% 叠氮化钠的磷酸盐缓冲盐水中以 1：10000 稀释。
Thiazole Orange 染色
1 网织红细胞分析^[1]
1)将全血 (5 μL) 加入稀释的染料溶液 (1 mL) 中，在 12 x 75 mm 的培养管中，在室温下孵育 1 小时。样本在 FACS IV，FACS 440 或 FACStar 上进行分析。
2）数据分析 (流式细胞术)：成熟红血球位于左侧和中间标记之间，网织红细胞位于中间和右侧标记之间。细胞类型的分配是通过人外周单核细胞混合的全血分析确定的。
2 DNA 电泳^[3]
1) 在缓冲液中混合琼脂糖 (约 1% w/v，具体百分比可根据所需的分离大小进行调整) (大约 70 mL 用于迷你凝胶 (8 x 7 cm))。常用的缓冲液为 TAE (Tris-醋酸-EDTA，40 mM Tris，20 mM 醋酸，1 mM EDTA，pH 约为 8.6) 或 TBE (Tris-硼酸-EDTA，90 mM Tris，90 mM硼酸，2 mM EDTA，pH 约为 8.3)。
2) 添加 Thiazole Orange，最终浓度为 1.3 μg/mL。
注意：在电泳后也可以用 Thiazole Orange 染色凝胶。
3) 微波加热琼脂糖，缓冲液和 Thiazole Orange 的混合物以溶解琼脂糖 (约 60 秒)。这个步骤通常被称为“融化”。如果需要，可以缓慢旋转 (5 秒) 以帮助溶解。让琼脂糖溶液固化成凝胶。
4) 将凝胶放入电泳仪中。使用加载染料加载 DNA 样本 (通常为 10 μL)。盖上盖子并连接电极。施加电压 (通常为迷你凝胶时约100V)，直到加载染料移动到适当的距离 (迷你凝胶约为 4-7 cm)。
注意：Thiazole Orange 带有正电荷，移动方向与电泳 DNA 相反。
5) 从电泳仪中取出凝胶，放在紫外线透视仪和蓝光透视仪上 (最大发射波长约为 470 nm)。
6) 在胶成像设备中选择合适的激发和发射设置。Thiazole Orange 的激发和发射 (λ_ex,max = 510 nm (488 nm 和 470 nm 也显示出强激发，此外在紫外线波长下也有强激发)；λ_em = 527 nm)。

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

476.61

C₂₆H₂₄N₂O₃S₂

107091-89-4

Room temperature in continental US; may vary elsewhere.

Please store the product under the recommended conditions in the Certificate of Analysis.

• [1]. Suss O, et al. Broad Applications of Thiazole Orange in Fluorescent Sensing of Biomolecules and Ions. Molecules. 2021 May 10;26(9):2828.  [Content Brief]

  [2]. Long W, et al. Molecular Recognition and Imaging of Human Telomeric G-Quadruplex DNA in Live Cells: A Systematic Advancement of Thiazole Orange Scaffold To Enhance Binding Specificity and Inhibition of Gene Expression. J Med Chem. 2021 Feb 25;64(4):2125-2138.  [Content Brief]

  [3]. Lu YJ, et al. Molecular Engineering of Thiazole Orange Dye: Change of Fluorescent Signaling from Universal to Specific upon Binding with Nucleic Acids in Bioassay. ACS Chem Biol. 2016 Apr 15;11(4):1019-29.  [Content Brief]

  [4]. O'Neil CS, et al. DNA Electrophoresis Using Thiazole Orange Instead of Ethidium Bromide or Alternative Dyes. J Vis Exp. 2019 Mar 31;(145)  [Content Brief]

  [5]. Piotr Klimkowski, et al. Design of thiazole orange oligonucleotide probes for detection of DNA and RNA by fluorescence and duplex melting. Org Biomol Chem. 2019 Jun 28; 17(24): 5943–5950.  [Content Brief]

  [6]. Linda G. Le, et al. Thiazole Orange: A New Dye for Reticulocyte Analysis. cytometry 7:508-517 (1986)