BCECF is a pH-sensitive fluorescent dye and can be used to monitor cellular pH ratiometrically. BCECF allows measurements in the physiological pH range 6.0–8.0. Excitation ratio: 490/440 nm; Emission intensity: 535 nm.

Fully treated cells show hydrogenosomes with an electron-dense deposit which aggregates to a variable extent.The staining is seen in the interior of hydrogenosomes in some instances. It is also observed by microscopy that the K⁺/H⁺ ionophor nigericin does not inhibit hydrogenosomal loading with BCECF^[1].
The pH-sensitive fluorescent dyes to measure cytosolic pH.
1.Prepare a 2 to 20 mM stock solution of BCECF in DMSO.
2.Prepare a 5-50 µM BCECF dye-loading solution in buffer solutions (HHBS or PBS).
3. Add 1000 µL/well (6-well plate),100 µL/well (96-well plate) or 25 µL/well (384-well plate) BCECF dye-loading solution into the cell plate.
4. Incubate the dye-loading plate in a cell incubator for 30-60 minutes.
5. Wash and replace the dye-loading solution with buffers.
6. Run the pH assay by monitoring the fluorescence at E_x/E_m = 490/535 nm or 430/535 nm for ratio measurements.

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

520.44

C₅₄H₄₀O₂₂

85138-49-4

Room temperature in continental US; may vary elsewhere.

-20deg;C, protect from light

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

DMSO : 62.5 mg/mL (120.09 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)。-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。

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 (4.00 mM); Clear solution

  此方案可获得 ≥ 2.08 mg/mL (4.00 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

• [1]. Scott DA, et al. Analysis of the uptake of the fluorescent marker 2′,7′-bis-(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF) by hydrogenosomes in Trichomonas vaginalis. Eur J Cell Biol. 1998 Jun;76(2):139-45.

The cells are palced in a petri dish and BCECF present in trichomonads is fixed for electron microsopy. Controns without BCECF loading, without DAB treatment, and without UV illumination are run in parallel. Trichomonads remaine viable during the procedure, except that without BCECF they are dead and disrupted by 4 h. Then the cell are lysed. After lysis, BCECF is added to the lysate to 20 μM, and the lysate is incubated,followed by 3 washes in lysis buffer to remove free dye and resuspension of the pellet in the original volume^[1].

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

• [1]. Scott DA, et al. Analysis of the uptake of the fluorescent marker 2′,7′-bis-(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF) by hydrogenosomes in Trichomonas vaginalis. Eur J Cell Biol. 1998 Jun;76(2):139-45.