NucPE1 (Nuclear Peroxy Emerald 1) is a nuclear-localized fluorescent hydrogen peroxide that is specifically localized to cellular nuclei without appended targeting moieties.

NucPE1 features two major visible region absorptions (λ_abs=468 nm, ε=27,300 M^-1cm^-1; λ_abs=490 nm, ε=26,000 M^-1cm^-1) and a weak emission (λ_em=530 nm, Φ=0.117). Reaction of NucPE1 with H₂O₂ triggers a fluorescence increase upon its conversion to fluorophore NucPE1, which possesses one major absorption band at 505 nm (ε=19,100 M^-1cm^-1) with enhanced emission (λ_em=530 nm, Φ=0.626). NucPE1 selectively accumulates in the nuclei of a variety of mammalian cell lines as well as in whole model organisms like C. clegans, where it can respond to subcellular changes in H₂O₂ fluxes^[1].

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

NucPE1 maintains the ability to selectively target nuclei in vivo. NucPE1 imaging reveals a reduction in nuclear H₂O₂ levels in worms overexpressing sir-2.1 compared to wildtype congeners, supporting a link between this longevity-promoting sirtuin protein and enhanced regulation of nuclear ROS pools^[1].

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

483.36

C₂₉H₃₀BNO₅

1404091-23-1

Room temperature in continental US; may vary elsewhere.

4deg;C, protect from light, stored under nitrogen

*In solvent : -80deg;C, 6 months; -20deg;C, 1 month (protect from light, stored under nitrogen)

In Vitro:;

DMSO : 25 mg/mL (51.72 mM; Need ultrasonic)

*

请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。
储备液的保存方式和期限：-80°C, 6 months; -20°C, 1 month (protect from light, stored under nitrogen)。-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。

In Vivo:

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液，再依次添加助溶剂：

——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用； 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶

• 1.

  请依序添加每种溶剂：;10% DMSO ;; 40% PEG300 ;; 5% Tween-80 ;; 45% saline

  Solubility: ≥ 2.5 mg/mL (5.17 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (5.17 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀；向上述体系中加入50 μL Tween-80，混合均匀；然后继续加入 450 μL生理盐水定容至 1 mL。

  将 0.9 g 氯化钠，完全溶解于 100 mL ddH₂O 中，得到澄清透明的生理盐水溶液

• [1]. Stanicka J, et al. NADPH oxidase-generated hydrogen peroxide induces DNA damage in mutant FLT3-expressing leukemia cells. J Biol Chem. 2015 Apr 10;290(15):9348-61.

  [2]. Dickinson BC, et al. A nuclear-localized fluorescent hydrogen peroxide probe for monitoring sirtuin-mediated oxidative stress responses in vivo. Chem Biol. 2011 Aug 26;18(8):943-8.

Excitation of NucPE1-loaded cells at 514 nm is carried out with an Ar laser and emission is collected using a META detector between 522–554 nm. Excitation of Hoechst 33342 is carried out using a MaiTai two-photon laser at 780-nm pulses and emission is collected between 436–501 nm. All images in an experiment are collected simultaneously using identical microscope settings. Image analysis is performed in Image J. The background fluorescence is measured in a data set by selecting an ROI outside of the cells. This background number is then used to set the threshold of the image and thereby select for NucPE1 signal. The mean fluorescence intensity of this signal is then measured and utilized as the fluorescent signal for that particular image. The same threshold settings are utilized for all images in an experiment^[1]. The measurement of nuclear H₂O₂ is achieved using NucPE1. The cells are incubated for 45 min at 10 μM NucPE1 in the dark. The incubation is followed by ishing and analysis by flow cytometry as explained above. Mitochondrial ROS are measured using MitoSOX probe. The cells are incubated with 5 μM MitoSOX for 15 min in the dark. The incubation is followed by ishing and analysis by flow cytometry^[2]. Live imaging of the NucPE1 probe is carried out using excitation at 488 nm with an argon laser, and emission is collected using a META detector at about 520 nm. The Hoechst dye is incubated together with NucPE1. When multiple staining of NucPE1 and PO1 is performed, the multitracking mode of scanning is applied for acquisition of the images^[2].

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

• [1]. Stanicka J, et al. NADPH oxidase-generated hydrogen peroxide induces DNA damage in mutant FLT3-expressing leukemia cells. J Biol Chem. 2015 Apr 10;290(15):9348-61.

  [2]. Dickinson BC, et al. A nuclear-localized fluorescent hydrogen peroxide probe for monitoring sirtuin-mediated oxidative stress responses in vivo. Chem Biol. 2011 Aug 26;18(8):943-8.