多药耐药性检测试剂盒——监测三种ABC转运蛋白 EFLUXX-ID® multidrug resistance assay kit

多药耐药性检测试剂盒——监测三种ABC转运蛋白
EFLUXX-ID® multidrug resistance assay kit

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EFLUXX-ID® multidrug resistance assay kit多药耐药性检测试剂盒——监测三种ABC转运蛋白                              EFLUXX-ID® multidrug resistance assay kit

多药耐药性检测试剂盒——监测三种ABC转运蛋白

简单免洗检测,可同时监测3种耐药性相关的ABC转运蛋白

耐药性是一种因ATP结合盒式(ATP Binding Cassette,ABC)膜转运蛋白家族上调介导所产生的现象。

ABC转运蛋白的过度表达了加速有毒物质从细胞中的去除,例如化疗剂从肿瘤细胞中排出,或抗生素从耐药菌株中排出。

Enzo的EFLUXX-ID® 多药耐药性检测试剂盒可实现同时对三种临床相关的 ABC 转运蛋白的功能检测:包括MDR1(p-糖蛋白)、MRP1/2 和 BCRP。

 

◆特点

● 简单免洗的操作步骤,结果可在1小时内获得

● 可检测与3种ABC转运蛋白(MDR1(p-糖蛋白)、MRP1/2和BCRP)活性相关的耐药性

● 通过单一专用染料可定量检测活细胞中的多药耐药性(MDR)活性,并以MDR 活性因子(MAF)表征

● 试剂盒内含有三种已知的MDR1(p-糖蛋白)、MRP1/2和BCRP蛋白特异性抑制剂

● 可检测Calcein AM染料无法检测的BCRP蛋白活性

● 有绿色、金色两种荧光染料可供选择

● 两种染料均可与表达GFP的细胞系或其他CELLESTIAL® 染料同时使用


作用机制


 EFLUXX-ID®使用了一种疏水性非荧光化合物,这种化合物容易穿透细胞膜,经细胞内酯酶水解成亲水性荧光染料。除非EFLUXX-ID® 染料被泵出至细胞外,否则酯酶裂解的染料会一直存在于细胞内。因此,表现出耐药性的细胞将染料主动运输泵出胞外,会出现荧光减弱的现象。EFLUXX-ID® 检测是目前少有可同时监测三种主要的ABC转运蛋白并能够分析单泵活性的试剂盒。

  荧光团

  兼容的EFLUXX-ID® 试剂

  Cy 3 & Cy 5的偶联物、Pl、7-AAD、APC、Texas Red、

  RFP、YFP、TAMRA、UV和Violet 染料

  EFLUXX-ID® 绿色荧光染料

  GFP/eGFP、RFP、PI、7-AAD、APC、Cy 5的偶联物、

  Rhodamine   123、Texas Red、UV和Violet   染料

  EFLUXX-ID® 金色荧光染料

表1:EFLUXX-ID® 多药耐药性染料与其他荧光染料的兼容性

多药耐药性检测试剂盒——监测三种ABC转运蛋白                              EFLUXX-ID® multidrug resistance assay kit

图1:对三种主要ABC转运蛋白的活性进行检测。使用EFLUXX -ID Green(上)、Gold(中)或Calcein AM(下)染料,通过流式细胞术评估CHO K1细胞中ABC转运蛋白的活性。与未处理的细胞相比,用ABC转运蛋白特异性抑制剂(图中阴影部分)处理可诱导染料保留在细胞内(图中实线部分)。平均荧光强度(MFI)的差异表明了对应蛋白的活性,在图中通过多药耐药性活性因子值(MAF)进行表征。更高的MAF值表明EFLUXX-ID染料对抑制剂有着良好的特异性。而Calcein AM染料的低MAF值表明其(常用的MDR检测探针)无法检测BCRP活性。

多药耐药性检测试剂盒——监测三种ABC转运蛋白                              EFLUXX-ID® multidrug resistance assay kit

图2:使用EFLUXX-ID® Green和EFLUXX-ID® Gold染料在CHO K1细胞中评估已知抑制剂对ABC转运蛋白活性的结果分析。将细胞与试剂盒中含有的MDR普通抑制剂(左一)、转运蛋白特异性抑制剂在37°C下孵育5 min。随后在37°C下用指定的染料染色细胞30 min,并立即利用流式细胞术进行分析。所用抑制剂:5 µM环胞霉素 A(MDR普通抑制剂)、20 µM维拉帕米(P-gp特异性抑制剂)、0.05 mM MK-571(MRP特异性抑制剂)、0.05 mM新生霉素(BCRP特异性抑制剂)。

多药耐药性检测试剂盒——监测三种ABC转运蛋白                              EFLUXX-ID® multidrug resistance assay kit

图3:EFLUXX-ID® Green 490/514 nm ex/em和Gold 530/570 nm ex/em 试剂的光谱特性可与其他常见的荧光染料进行多重检测。

多药耐药性检测试剂盒——监测三种ABC转运蛋白                              EFLUXX-ID® multidrug resistance assay kit

◆产品详情


  应用

  流式细胞术,荧光显微镜

  应用说明

  EFLUXX-ID® 多药耐药性检测试剂盒可对活细胞(悬浮细胞和贴壁细胞)中的耐药表型进行检测和分析。

  品质保证

  每批次EFLUXX-ID® Gold耐药检测试剂盒的样品试剂盒均已按照操作手册中的步骤,对第5代和第20代间的

  CHO K1细胞系进行染色。获得的结果如下

  1.   维拉帕米介导的抑制后MAF>80

  2. MK-571介导的抑制后MAF>60

  3. 新生霉素介导的抑制后MAF>40

  包装

  100 assays(流式细胞术)

  使用/稳定性

  自交货后,试剂盒组分在适当的储存条件下可稳定保存一年。

  储存条件

  避光。避免反复冻融。

  运输条件

  蓝冰运输

  短期储存

  -20°C

  长期储存

  -80°C

  组分

  2 瓶,EFLUXX-ID® Gold或EFLUXX-ID® Green检测试

  300 nmoles,MDR1 抑制剂(维拉帕米)

  750 nmoles,MRP   抑制剂(MK-571)

  1.5 µmoles,BCRP 抑制剂(新生霉素)

  500 µL,碘化丙啶

  科学背景

  耐药性是一种由跨膜ATP结合盒(ATP   Binding Cassette   ,简称ABC)转运蛋白家族上调介导的现象。

  ABC转运蛋白的过表达加速了毒性剂从细胞中去除,例如化疗剂从肿瘤细胞中流出,或抗生素从耐药菌株中流出。

  监管状态

  RUO – 仅供研究用

参考文献



◆产品参考文献


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Exploiting off-target effects of estrogen deprivation to sensitize estrogen receptor negative breast cancer to immune killing: B. Wolfson, et al.; J. Immunother. Cancer 9, 2258 (2021), 摘要;


 2.

MicroRNA-324-5p regulates stemness, pathogenesis and sensitivity to bortezomib in multiple myeloma cells by targeting hedgehog signaling: B. Tang, et al.; Int. J. Cancer 142, 109 (2018), Application(s): Flow Cytometry; multiple myeloma, 摘要Full Text


 3.

Upregulation of FOXM1 in a subset of relapsed myeloma results in poor outcome: C. Gu, et al.; Blood Cancer J. 8, 22 (2018), Application(s): Flow Cytometry; myeloma cells, 摘要Full Text


 4.

Resistin induces multidrug resistance in myeloma by inhibiting cell death and upregulating ABC transporter expression: J. Pang, et al.; Haematologica 102, 1273 (2017), Application(s): Flow Cytometry; myeloma cells, 摘要Full Text


 5.

Hepatocyte SLAMF3 reduced specifically the multidrugs resistance protein MRP-1 and increases HCC cells sensitization to anti-cancer drugs: G. Fouquet, et al.; Oncotarget 7, 32493 (2016), 摘要; Full Text


 6.

Intercellular transfer of P-glycoprotein in human blood-brain barrier endothelial cells is increased by histone deacetylase inhibitors: A. Noack, et al.; Sci. Rep. 6, 29253 (2016), Application(s): Flow cytometry of Pgp-EGFP transfer in co-cultured cells, 摘要Full Text


 7.

The influence of a caveolin-1 mutant on the function of P-glycoprotein: C.Y. Lee, et al.; Sci. Rep. 6, 20486 (2016), 摘要Full Text


 8.

RhoGDI2 up-regulates P-glycoprotein expression via Rac1 in gastric cancer cells: Z. Zheng, et al.; Cancer Cell Int. 15, 41 (2015), Application(s): Monitoring of MDR1 functionality by flow cytometry, 摘要Full Text


 9.

Role of NEK2A in human cancer and its therapeutic potentials: J. Xia, et al.; Biomed. Res. Int. 2015, 862461 (2015), 摘要; Full Text

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Sonic hedgehog-glioma associated oncogene homolog 1 signaling enhances drug resistance in CD44+/Musashi-1+ gastric cancer stem cells: M. Xu, et al.; Cancer Lett. 369, 124 (2015), 摘要;


11.

Drug-induced trafficking of p-glycoprotein in human brain capillary endothelial cells as demonstrated by exposure to mitomycin C: A. Noack, et al.; PLoS One 9, e88154 (2014), Application(s): MDR status by flow cytometry in microvascular endothelial cells, 摘要全文


12.

NEK2 induces drug resistance mainly through activation of efflux drug pumps and is associated with poor prognosis in myeloma and other cancers: W. Zhou, et al.; Cancer Cell 23, 48 (2013), Application(s): Dye Efflux Assay for Multidrug Resistance was performed with ARP1 cancer cell lines and MCF7 cells using Gold detection reagent on flow cytometry, 摘要;


13.

Localization microscopy (SPDM) reveals clustered formations of P-glycoprotein in a human blood-brain barrier model: O. Huber, et al.; PLoS One 7, e44776 (2012), Application(s): MDR status by flow cytometry in immortalized human brain endothelial cells, 摘要全文


14.

Sensitive and specific fluorescent probes for functional analysis of the three major types of mammalian ABC transporters.: I. Lebedeva, et al.; PLoS One 6, e22429 (2011), 摘要全文

◆其他文献


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P-Glycoprotein-mediated resistance to Hsp90-directed therapy is eclipsed by the heat shock response: A.K. McCollum, et al.; Cancer Res. 68, 7419 (2008), 摘要;


 2.

Glutathione export during apoptosis requires functional multidrug resistance-associated proteins: C.L. Hammond, et al.; J. Biol. Chem. 282, 14337 (2007), 摘要;


 3.

From MDR to MXR: new understanding of multidrug resistance systems, their properties and clinical significance: T. Litman, et al.; Cell Mol. Life Sci. 58, 931 (2001), 摘要;


 4.

Increased levels of the multidrug resistance protein in lateral membranes of proliferating hepatocyte-derived cells: H. Roelofsen, et al.; Gastroenterology 112, 511 (1997), 摘要;


 5.

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Polarized efflux of 2’,7’-bis(2-carboxyethyl)-5(6)-carboxyfluorescein from cultured epithelial cell monolayers: G.K. Collington, et al.; Biochem. Pharmacol. 44, 417 (1992), 摘要;


11.

Epithelial secretion of vinblastine by human intestinal adenocarcinoma cell (HCT-8 and T84) layers expressing P-glycoprotein: J. Hunter, et al.; Br. J. Cancer 64, 437 (1991), 摘要;


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产品列表
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ENZ-51029-K100 EFLUXX-ID® Green multidrug resistance assay kit
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