Hoechst 33258 is afluorescent dyethat emits blue fluorescence when bound to dsDNA.

IC50: 51.31±4.56 μM (HeLa cell), 32.43±3.27 μM (HL60 cell), 15.42 ± 2.16 μM (U937 cell)^[1]

Hoechst 33258, a fluorescent compound with a head-to-tail bis-benzimidazole structure, is initially found to be cytotoxic against L1210 murine leukemia. Hoechst 33258 is evaluated for their cytotoxicity against human tumor cell lines, which are cervix carcinoma cell line (HeLa), Human promyelocytic leukemia cell (HL60) and U937 cell Line. The IC₅₀determined in the case of HeLa, HL60 and U937 is 51.31±4.56, 32.43±3.27 and 15.42±2.16 μM for Hoechst 33258, respectively^[1]. The cytotoxic property of Hoechst 33258 is investigated on a panel of seven tumour cell lines of different histological origin and Madine-Darby canine kidney (MDCK) normal cells. All cell lines, except MCF-7, exposed to Hoechst 33258 exhibit GI₅₀from 84×10^-6to 191.5×10^-6mol/dm³. Under the same experimental conditions, Hoechst 33258, used as a binder reference compound, stops the cell cycle in S phase and G0/G1^[2].

424.50

Solid

C₂₅H₂₄N₆O

23491-44-3

Room temperature in continental US; may vary elsewhere.

-20°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

DMSO : 41.67 mg/mL(98.16 mM;ultrasonic and warming and heat to 60℃)

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℃, 6 months; -20℃, 1 month (protect from light)。-80℃ 储存时，请在 6 个月内使用，-20℃ 储存时，请在 1 个月内使用。

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: ≥ 2.08 mg/mL (4.90 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (4.90 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 20.8 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
• 2.

  请依序添加每种溶剂： 10% DMSO    90% (20%SBE-β-CDin saline)

  Solubility: ≥ 2.08 mg/mL (4.90 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (4.90 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 20.8 mg/mL 的澄清 DMSO 储备液加到 900 μL20% 的 SBE-β-CD 生理盐水水溶液中，混合均匀。

  将 2 g 磺丁基醚 β-环糊精加入 5 mL 生理盐水中，再用生理盐水定容至 10 mL，完全溶解，澄清透明

• 
  Description: Hoechst 33258 can be used for detecting nuclei.
  Method: For cell staining.
  1. Fix, penetrate and seal cells first.
  2. Incubate cells with Hoechst 33258 (1:1000; 30 min; dark).
  3. Wash cells with PBS for five times and use a fluorescence microscope (ECLIPSE TE2000-5) for image.
• 
  Description: Hoechst 33258 can be used for detecting nuclei and staining nuclei of live and dead (apoptotic/necrotic).
  Method: For cell staining.
  1. Incubate cultured cells with 1 mL of the solution prepared with Hoechst 33258 (7.5 μg/mL; 37℃; 5% CO₂; 30 min; 1×PBS).
  2. Use a Evos FLc Fluorescent Inverted Microscope for image.
• 
  Description: Hoechst 33258 can be used for detecting nuclei, it can be used in apoptosis detection assays.
  Method: For cell staining.
  1. Fix cells in paraformaldehyde after transfection and wash cells with PBS.
  2. Stain cells with Hoechst 33258 (30 min; 20℃; dark).
  3. Use a fluorescence microscope (Leica, Solms, Germany) for image.
• 
  Description: Hoechst 33258 can be used to visualize the nuclear DNA.