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CFDA-SE (Synonyms: 5(6)Carboxyfluorescein diacetate succinimidyl ester, 5(6)-CFDA N-succinmidyl ester)

Catalog No.GC14056

CFDA-SE는 세포막 투과성을 가진 카르복시세인 디아세테이트, 스크시니미딜 에스터의 전체 이름으로, 생존 가능한 세포 추적 또는 세포 증식 감지에 널리 사용되는 형광 염료입니다.

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CFDA-SE Chemical Structure

Cas No.: 150347-59-4

Size 가격 재고 수량
10mg
US$40.00
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25mg
US$84.00
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50mg
US$164.00
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100mg
US$339.00
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Sample solution is provided at 25 µL, 10mM.

Product has been cited by 1 publications

Product Documents

Quality Control & SDS

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Protocol

Protocol for Cell labeling and counting with CFDA-SE [1]:

Cells were counted and stained with CFDA-SE, as follows:

  1. 10 mm stock solution of CFDA-SE (Molecular Probes) was prepared in DMSO and aliquots were stored at -20°C in the presence of a desiccant.
  2. Harvested cells were washed in HBSS and FBS 1%, resuspended at 20 × 106/mL HBSS with FBS 1%, and stained in a 1:2000 dilution of stock CFDA-SE in HBSS to achieve a final concentration of 2.5 μM (cells were mixed to CFDA-SE 1:1 v/v). The procedure was carried out at room temperature in the dark.
  3. At the end of the incubation period (8 min) the reaction was stopped by adding an equal volume of FBS. Cells were gently pelleted at 300 g for 10 min and washed (400 g for 10 min) twice in culture medium
  4. Cells were plated and the day after the first flow cytometric acquisition was performed (time point 1). In populations showing a particularly high proliferative rate, two or more generations of cells might be already evident at this point. However, the analysis at the subsequent time points was carried out by comparing the stain distribution with the one described at time point 1. Dead cells were excluded and analysis of acquisition files was performed separately on the two subpopulations.

 

This protocol only provides a guideline, and should be modified according to your specific needs.

References:

[1]. Urbani S, Caporale R,et,al. Use of CFDA-SE for evaluating the in vitro proliferation pattern of human mesenchymal stem cells. Cytotherapy. 2006;8(3):243-53. doi: 10.1080/14653240600735834. PMID: 16793733.

Cell experiment [1]:

Cell lines

Human erythroleukaemic cell line K562, mouse lymphoma cell line YAC-1, human mammary cancer cell line MCF-7 and human melanoma cell line A375

Preparation method

The 5 mM CFDA-SE stock in DMSO was diluted to different concentrations (2 μM, 3 μM, 4 μM, 5 μM, 10 μM and 20 μM) in PBS with a total volume of 1 ml. Cells were added to equal volume of CFDA-SE with different concentrations and incubated at 37 C for 5, 6, 7, 8, 10 and 15 min with agitation.

Reaction Conditions

1, 1.5, 2, 2.5, 5 and 10 µM CFDA-SE

Applications

CFDA-SE at 2.5 μM stained more than 95% of the cells on all cell lines tested, and dose-dependently increased fluorescence intensity of stained cells. The optimal concentration for K562 and YAC-1 was found to be 2.5 μM, while the optimal concentrations for A375 and MCF-7 were found to be 5 μM and 10 μM, respectively. Within the 6 h experiment period, no cytotoxicity related to CFDA-SE was observed.

Animal experiment [2]:

Animal models

C57BL/6 mice, aged 5 ~ 8 weeks

Preparation method

 Injected CFDA-SE into thymic lobe

Dosage form

The concentration of CFDA-SE is 10 μM

Applications

CFDA-SE, at 80 times the concentration used for in vitro labeling, was nontoxic and labeled randomly approximately 15% of thymocytes 24 h after injection. The turnover rate of labeled thymic emigrants in the lymph nodes was in the order of 21 days. Thus, CFDA-SE may serve as a powerful tool in relatively long-term migration studies.

References:

[1]: Wang XQ, Duan XM,et,al. Carboxyfluorescein diacetate succinimidyl ester fluorescent dye for cell labeling. Acta Biochim Biophys Sin (Shanghai). 2005 Jun;37(6):379-85. doi: 10.1111/j.1745-7270.2005.00051.x. PMID: 15944752.

[2]: Graziano M, St-Pierre Y, et,al. The fate of thymocytes labeled in vivo with CFSE. Exp Cell Res. 1998 Apr 10;240(1):75-85. doi: 10.1006/excr.1997.3900. PMID: 9570923.

Background

CFDA-SE는 세포막 투과성을 가진 카르복시세인 디아세트산, 스크시니미딜 에스터의 약칭으로, 생존 가능한 세포 추적 또는 세포 증식 감지에 널리 사용되는 형광 염료입니다[1]. CFDA-SE로 표지된 세포의 형광은 매우 균일하고 안정적이며, 형광은 두 개의 후손 세포에 고르게 분배될 수 있습니다[2]. CFDA-SE는 강력한 초록색 형광을 방출할 수 있으며, Ex=494 nm, Em=521nm입니다.

인간 적혈구 백혈병 세포주 K562 및 다른 세포주에서는 모든 세포가 CFDA-SE에 의해 상대적으로 높은 형광 강도로 표시되지만, 표시의 효율성은 매우 가변적입니다. 전자밀도 소포가 표시된 세포에서 초미세 구조 수준에서 관찰됩니다. 형광 현미경으로 볼 때 전체 세포에 이산화물 라벨링이 가능합니다. 전체 세포는 6 주 동안 좋은 라벨을 유지합니다. CFDA-SE 라벨링은 비교적 쉽고, 세포에 대한 비독성이며 방사능이 아닙니다.

CFDA-SE 방법을 사용한 체외 림프구 세포 증식 분석은 아직 분류되지 않은 PID 환자의 유전학적 분석을 위한 미토젠성 T 림프구 반응 평가에 신뢰성 있고 실용적인 선택이다. 플로우 사이트메트리에서 NM이 감지될 때, FL1 검출 채널 CFDA-SE -라벨링된 세포를 채용할 수 있으며 형광 현미경도 인접한 세포를 염색하지 않고 CFDA-SE -라벨링된 세포를 관찰하는 데 사용될 수 있다.

CFDA-SE는 CD34+ 세포에 라벨링하는 데 효과적이라고 보고되었습니다. 태아 간이나 흉선에서 분리한 CD34+ 세포를 CFDA-SE로 라벨링하고 RAG-2/IL-2Rγ/ 마우스의 피하에 이식된 인간 흉선 내부에 주입했습니다. 1~4주 후, CFDA-SE 라벨은 T세포뿐만 아니라 CD123+/high CD4+CD45RA+ pDC2에서도 발견되어 CD34+ 세포가 흉선 내부에서 pDC2로 발전할 수 있다는 것을 나타내었습니다. pDC2 외에도, 세포면 마커인 CD11c, CD83 및 CD80으로 결정된 성숙한 형질을 가진 CFDA-SE 라벨의 수지상 대조 그래프내에서 양성 반응을 보인 수지상 대조 그래프내 주입된 인간 흉선 내 dendritic cell들이 발견되었습니다. 인간 흉선 이식체를 운반하는 마우스의 말단 부위에서는 pDC2가 발견되지 않았으므로, thymus 내부의 pDC2가 thymus 밖으로 이동하지 못한다는 것을 나타냅니다. C57BL/6 마우스에서는 CFDA-SE가 vitro 라벨링에 사용된 농도의 80배로 주입해도 비독성이며, 주사 후 24시간 이내에 thymocyte의 약 15%를 임의로 라벨링합니다. 라벨링된 흉선 이동세포의 회전율은 약 21일입니다. 따라서 CFDA-SE는 상대적으로 장기간 이동 연구에 강력한 도구로 사용될 수 있습니다.

References:
[1]: Chen JC, Chang ML, et,al. A kinetic study of the murine mixed lymphocyte reaction by 5,6-carboxyfluorescein diacetate succinimidyl ester labeling. J Immunol Methods. 2003 Aug;279(1-2):123-33. doi: 10.1016/s0022-1759(03)00236-9. PMID: 12969553. [2]: Lyons AB, Blake SJ, et,al. Flow cytometric analysis of cell division by dilution of CFSE and related dyes. Curr Protoc Cytom. 2013;Chapter 9:Unit9.11. doi: 10.1002/0471142956.cy0911s64. PMID: 23546777. [3]: Wang XQ, Duan XM, et,al. Carboxyfluorescein diacetate succinimidyl ester fluorescent dye for cell labeling. Acta Biochim Biophys Sin (Shanghai). 2005 Jun;37(6):379-85. doi: 10.1111/j.1745-7270.2005.00051.x. PMID: 15944752. [4]: Gruber HE, Leslie KP, et,al. Optimization of 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester for labeling human intervertebral disc cells in vitro. Biotech Histochem. 2000 May;75(3):118-23. doi: 10.3109/10520290009066489. PMID: 10950173. [5]: Azarsiz E, Karaca N, et,al. In vitro T lymphocyte proliferation by carboxyfluorescein diacetate succinimidyl ester method is helpful in diagnosing and managing primary immunodeficiencies. J Clin Lab Anal. 2018 Jan;32(1):e22216. doi: 10.1002/jcla.22216. Epub 2017 Apr 6. PMID: 28383134; PMCID: PMC6816938. [6]: Li X, Dancausse H, et,al. Labeling Schwann cells with CFSE-an in vitro and in vivo study. J Neurosci Methods. 2003 May 30;125(1-2):83-91. doi: 10.1016/s0165-0270(03)00044-x. PMID: 12763234. [7]: Weijer K, Uittenbogaart CH, et,al. Intrathymic and extrathymic development of human plasmacytoid dendritic cell precursors in vivo. Blood. 2002 Apr 15;99(8):2752-9. doi: 10.1182/blood.v99.8.2752. PMID: 11929763. [8]: Graziano M, St-Pierre Y, et,al. The fate of thymocytes labeled in vivo with CFSE. Exp Cell Res. 1998 Apr 10;240(1):75-85. doi: 10.1006/excr.1997.3900. PMID: 9570923.

Chemical Properties

Cas No. 150347-59-4 SDF
Synonyms 5(6)Carboxyfluorescein diacetate succinimidyl ester, 5(6)-CFDA N-succinmidyl ester
Chemical Name 5-(((2,5-dioxopyrrolidin-1-yl)oxy)carbonyl)-3-oxo-3H-spiro[isobenzofuran-1,9'-xanthene]-3',6'-diyl diacetate
Canonical SMILES O=C1N(OC(C2=CC=C(C3(C(C=CC(OC(C)=O)=C4)=C4OC5=C3C=CC(OC(C)=O)=C5)OC6=O)C6=C2)=O)C(CC1)=O
Formula C29H19NO11 M.Wt 557.46
Solubility ≥ 37.2mg/mL in DMSO with ultrasonic Storage Store at -20°C, protect from light
General tips Please select the appropriate solvent to prepare the stock solution according to the solubility of the product in different solvents; once the solution is prepared, please store it in separate packages to avoid product failure caused by repeated freezing and thawing.Storage method and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored at -20°C, please use it within 1 month.
To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time.
Shipping Condition Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request.

Complete Stock Solution Preparation Table

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1 mg 5 mg 10 mg
1 mM 1.7939 mL 8.9693 mL 17.9385 mL
5 mM 0.3588 mL 1.7939 mL 3.5877 mL
10 mM 0.1794 mL 0.8969 mL 1.7939 mL
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