JC-1线粒体膜电位检测试剂盒（JC-1 Mitochondrial Membrane Potential Assay Kit）是一种以JC-1 为荧光探针，快速灵敏地检测细胞、组织或纯化的线粒体膜电位变化的试剂盒，可以用于早期的细胞凋亡检测，也是用来检测细胞早期凋亡的常用方法。
JC-1 是一种广泛用于检测线粒体膜电位△Ψm 的理想荧光探针，JC-1染料以电势依赖性的方式积聚在线粒体内。正常线粒体内，JC-1聚集在线粒体基质中形成聚合物，聚合物发出强烈的红色荧光（Ex=585 nm, Em=590 nm）；不健康的线粒体由于膜电位的下降或丧失，JC-1只能以单体的形式存在于胞浆中，产生绿色荧光（Ex=514 nm, Em=529 nm）。JC-1不仅可用于定性检测，因颜色的变化可以非常直接的反映出线粒体膜电位的变化。也可以用于定量检测，因线粒体的去极化程度可以通过红/绿荧光强度的比例来衡量。观察时，只需使用常规的观察红色荧光和绿色荧光的设置即可。
线粒体膜电势的破坏是细胞凋亡早期发生的一个标志性事件。细胞受到凋亡诱导后线粒体膜电位的变化使得膜的通透性发生改变。膜通透性的增加，使得线粒体蛋白包括细胞色素C、Smac/DIABLO、HtrA2/OMI、核酸内切酶G（EndoG）、凋亡诱导因子（AIF）等从线粒体基质释放到细胞浆。细胞色素C的释放伴随膜电位的完全丧失，进而引发细胞凋亡酶的级联效应。
本试剂盒提供了CCCP 作为诱导线粒体膜电位下降的阳性对照。对于六孔板中的样品，本试剂盒共可以检测100 个样品；对于12 孔中的样品，本试剂盒共可以检测200 个样品。
冰袋（wet ice）运输。
-20℃避光保存，尽量避免反复冻融，有效期一年。超纯水和JC-1 染色缓冲液（5×）也可4℃保存。
Q：JC-1 探针和试剂盒有什么区别，客户做细胞实验用哪一款？
A:前者只是一个探针的粉末；后者是液体，又多了缓冲液，阳性对照和超纯水。两个都可以。
Q：JC-1 试剂盒检测的时候，客户要用流式检测，是要先把贴壁细胞消化下来再按照悬浮细胞处理吗？还是先染完色再消化下来去检测？
A:先消化下来制成细胞悬液再染色。
Q：阴性对照中线粒体都是正常的线粒体，为什么也可以观察到绿色的荧光，而且红色的也看不到细胞形态的原因是？
A：阴性对照可以拍到绿色可能是因为染色的时候加入的染料较多，从而使阴性对照可以排除绿色的荧光，可以适当的降低探针的浓度。绿色可以看到整个形态是因为单体（绿光）是散落在细胞质中的，而聚合体（红色）是结合到线粒体上的，有些细胞中线粒体分布是不均匀的，能否完整的看到细胞的形态是和线粒体的分布也是有一定的原因的。
Q：石蜡切片和冰冻切片可以做JC-1吗？
A：不可以，JC-1实验的样本必须是活细胞。
Q：JC-1孵育完成后不能及时检测，可以固定后再检测吗？
A：不可以，JC-1用于活细胞样本的检测，固定后细胞死亡；此外，染色后放置过久会导致荧光淬灭，建议染色30 min内完成检测。
Q：组织样本可不可以用JC-1检测凋亡？
A：无法直接检测组织，可先将组织制备成单细胞悬液，再按照悬浮细胞的操作步骤来检测，需对制备方案进行摸索。
Q：JC-1工作液的配置中出现绿色沉淀应如何处理？
A：取适量JC-1（200×），按照每50μL JC-1（200×）加入8 mL 超纯水的比例稀释JC-1，剧烈震荡充分溶解并混匀JC-1，然后再加入2 mL JC-1 染色缓冲液（5×），如溶液后存在绿色不溶沉淀可加入千分之一的Tween 20。
Q：该JC-1是否可用于分离纯化的完整线粒体膜电位的检测？JC-1的建议工作液如何配置？对应的纯化线粒体的量应为多少？加入JC-1后多久可以检测？
A：可以用于纯化分离得到的完整线粒体的膜电位的检测。
JC-1的工作液及线粒体用量的比例为：0.9 mL JC-1染色工作液（1×）中加入0.1 mL总蛋白量为10-100 µg纯化的线粒体。
加入JC-1工作液3-5 min后即可进行检测。
但需留意对于冷冻的纯化线粒体不可用于进行膜电位的检测，因线粒体膜经冻融被破坏。
[1] Chen LJ, Zhang NN, Zhou CX, et al. Gm364 coordinates MIB2/DLL3/Notch2 to regulate female fertility through AKT activation. Cell Death Differ. 2022;29(2):366-380. doi:10.1038/s41418-021-00861-5(IF:15.828)
[2] Li H, Zhu J, Xu YW, et al. Notoginsenoside R1-loaded mesoporous silica nanoparticles targeting the site of injury through inflammatory cells improves heart repair after myocardial infarction. Redox Biol. 2022;54:102384. doi:10.1016/j.redox.2022.102384(IF:11.799)
[3] Ding M, Ning J, Feng N, et al. Dynamin-related protein 1-mediated mitochondrial fission contributes to post-traumatic cardiac dysfunction in rats and the protective effect of melatonin. J Pineal Res. 2018;64(1):10.1111/jpi.12447. doi:10.1111/jpi.12447(IF:10.391)
[4] Wang D, Zhang L, Hu A, et al. Loss of 4.1N in epithelial ovarian cancer results in EMT and matrix-detached cell death resistance. Protein Cell. 2021;12(2):107-127. doi:10.1007/s13238-020-00723-9(IF:10.164)
[5] Chen P, Hu T, Liang Y, et al. Neddylation Inhibition Activates the Extrinsic Apoptosis Pathway through ATF4-CHOP-DR5 Axis in Human Esophageal Cancer Cells. Clin Cancer Res. 2016;22(16):4145-4157. doi:10.1158/1078-0432.CCR-15-2254(IF:8.738)
[6] Chen YJ, Hong WF, Liu ML, et al. An integrated bioinformatic investigation of mitochondrial solute carrier family 25 (SLC25) in colon cancer followed by preliminary validation of member 5 (SLC25A5) in tumorigenesis. Cell Death Dis. 2022;13(3):237. Published 2022 Mar 14. doi:10.1038/s41419-022-04692-1(IF:8.469)
[7] Li G, Li Z, Yang Z, Leshem Y, Shen Y, Men S. Mitochondrial heat-shock cognate protein 70 contributes to auxin-mediated embryo development. Plant Physiol. 2021;186(2):1101-1121. doi:10.1093/plphys/kiab138(IF:8.340)
[8] Chen LJ, Yang ZX, Wang Y, et al. Single xenotransplant of rat brown adipose tissue prolonged the ovarian lifespan of aging mice by improving follicle survival. Aging Cell. 2019;18(6):e13024. doi:10.1111/acel.13024(IF:7.346)
[9] Chen P, Hu T, Liang Y, et al. Synergistic inhibition of autophagy and neddylation pathways as a novel therapeutic approach for targeting liver cancer. Oncotarget. 2015;6(11):9002-9017. doi:10.18632/oncotarget.3282(IF:6.359)
[10] Zhou R, Xia M, Zhang L, et al. Fine particles in surgical smoke affect embryonic cardiomyocyte differentiation through oxidative stress and mitophagy. Ecotoxicol Environ Saf. 2021;217:112259. doi:10.1016/j.ecoenv.2021.112259(IF:6.291)
[11] Zhao M, Gu L, Li Y, et al. Chitooligosaccharides display anti-tumor effects against human cervical cancer cells via the apoptotic and autophagic pathways. Carbohydr Polym. 2019;224:115171. doi:10.1016/j.carbpol.2019.115171(IF:6.044)
[12] Chen S, Dong G, Wu S, Liu N, Zhang W, Sheng C. Novel fluorescent probes of 10-hydroxyevodiamine: autophagy and apoptosis-inducing anticancer mechanisms. Acta Pharm Sin B. 2019;9(1):144-156. doi:10.1016/j.apsb.2018.08.003(IF:6.014)
[13] Huang CX, Jiang ZX, Du DY, Zhang ZM, Liu Y, Li YT. The MFF-SIRT1/3 axis, regulated by miR-340-5p, restores mitochondrial homeostasis of hypoxia-induced pulmonary artery smooth muscle cells. Lab Invest. 2022;102(5):515-523. doi:10.1038/s41374-022-00730-w(IF:5.662)
[14] Li YR, Peng RR, Gao WY, et al. The ubiquitin ligase KBTBD8 regulates PKM1 levels via Erk1/2 and Aurora A to ensure oocyte quality. Aging (Albany NY). 2019;11(4):1110-1128. doi:10.18632/aging.101802(IF:5.515)
[15] Zhou LJ, Mo YB, Bu X, et al. Erinacine Facilitates the Opening of the Mitochondrial Permeability Transition Pore Through the Inhibition of the PI3K/ Akt/GSK-3β Signaling Pathway in Human Hepatocellular Carcinoma [retracted in: Cell Physiol Biochem. 2021;55(5):674]. Cell Physiol Biochem. 2018;50(3):851-867. doi:10.1159/000494472(IF:5.500)
[16] Chen C, Ma J, Xu Z, et al. Rosmarinic Acid Inhibits Platelet Aggregation and Neointimal Hyperplasia In Vivo and Vascular Smooth Muscle Cell Dedifferentiation, Proliferation, and Migration In Vitro via Activation of the Keap1-Nrf2-ARE Antioxidant System. J Agric Food Chem. 2022;70(24):7420-7440. doi:10.1021/acs.jafc.2c01176(IF:5.279)
[17] Chen A, An Y, Huang W, et al. Highly Water-Preserving Zwitterionic Betaine-Incorporated Collagen Sponges With Anti-oxidation and Anti-inflammation for Wound Regeneration. Front Cell Dev Biol. 2020;8:491. Published 2020 Jul 15. doi:10.3389/fcell.2020.00491(IF:5.186)
[18] Yao Y, Cui L, Ye J, et al. Dioscin facilitates ROS-induced apoptosis via the p38-MAPK/HSP27-mediated pathways in lung squamous cell carcinoma. Int J Biol Sci. 2020;16(15):2883-2894. Published 2020 Sep 2. doi:10.7150/ijbs.45710(IF:4.858)
[19] Sun Y, Sha B, Huang W, et al. ML323, a USP1 inhibitor triggers cell cycle arrest, apoptosis and autophagy in esophageal squamous cell carcinoma cells. Apoptosis. 2022;27(7-8):545-560. doi:10.1007/s10495-022-01736-x(IF:4.677)
[20] Lu H, Feng Y, Hu Y, et al. Spondin 2 promotes the proliferation, migration and invasion of gastric cancer cells. J Cell Mol Med. 2020;24(1):98-113. doi:10.1111/jcmm.14618(IF:4.658)
[21] Wu X, Yu H, Zhou H, et al. Proanthocyanidin B2 inhibits proliferation and induces apoptosis of osteosarcoma cells by suppressing the PI3K/AKT pathway. J Cell Mol Med. 2020;24(20):11960-11971. doi:10.1111/jcmm.15818(IF:4.486)
[22] Du L, Wang Y, Li CR, et al. Rat BAT xenotransplantation recovers the fertility and metabolic health of PCOS mice. J Endocrinol. 2021;248(2):249-264. doi:10.1530/JOE-20-0068(IF:4.286)
[23] Liu Q, Tao B, Liu G, et al. Thromboxane A2 Receptor Inhibition Suppresses Multiple Myeloma Cell Proliferation by Inducing p38/c-Jun N-terminal Kinase (JNK) Mitogen-activated Protein Kinase (MAPK)-mediated G2/M Progression Delay and Cell Apoptosis. J Biol Chem. 2016;291(9):4779-4792. doi:10.1074/jbc.M115.683052(IF:4.258)
[24] Zhou CX, Wang Y, Shi LY, et al. GTPases Arf5 and Arl2 function partially distinctly during oocyte meiosis. J Cell Biochem. 2021;122(2):198-208. doi:10.1002/jcb.29839(IF:4.237)
[25] Zhang H, Xu Y, Deng G, et al. SAA1 knockdown promotes the apoptosis of glioblastoma cells via downregulation of AKT signaling. J Cancer. 2021;12(9):2756-2767. Published 2021 Mar 10. doi:10.7150/jca.48419(IF:4.207)
[26] Liang WL , Wen Y , Huang F , et al. Chrysanthemum ethanol extract induced loss of Kupffer cells via the mitochondria-dependent apoptotic pathway. Food Funct. 2020;11(10):8866-8877. doi:10.1039/d0fo00695e(IF:4.171)
[27] Liu S, Xu DS, Ma JL, Huang P, Wu D, Ren LQ. LncRNA H19 Mitigates Oxidized Low-Density Lipoprotein Induced Pyroptosis via Caspase-1 in Raw 264.7 Cells. Inflammation. 2021;44(6):2407-2418. doi:10.1007/s10753-021-01511-1(IF:4.092)
[28] Sha B, Chen X, Wu H, et al. Deubiquitylatinase inhibitor b-AP15 induces c-Myc-Noxa-mediated apoptosis in esophageal squamous cell carcinoma. Apoptosis. 2019;24(9-10):826-836. doi:10.1007/s10495-019-01561-9(IF:4.021)
[29] Chen M, Yang J, Li L, et al. Metabolomics Reveals that Cysteine Metabolism Plays a Role in Celastrol-Induced Mitochondrial Apoptosis in HL-60 and NB-4 Cells. Sci Rep. 2020;10(1):471. Published 2020 Jan 16. doi:10.1038/s41598-019-57312-y(IF:3.998)
[30] Hu T, Zhang J, Sha B, et al. Targeting the overexpressed USP7 inhibits esophageal squamous cell carcinoma cell growth by inducing NOXA-mediated apoptosis. Mol Carcinog. 2019;58(1):42-54. doi:10.1002/mc.22905(IF:3.851)
[31] Zhang X, Wu W, Luo Y, Wang Z. Transcranial Photobiomodulation Therapy Ameliorates Perioperative Neurocognitive Disorder Through Modulation of Mitochondrial Function in Aged Mice. Neuroscience. 2022;490:236-249. doi:10.1016/j.neuroscience.2021.12.033(IF:3.590)
[32] Liu JZ, Hu YL, Feng Y, et al. Rafoxanide promotes apoptosis and autophagy of gastric cancer cells by suppressing PI3K /Akt/mTOR pathway. Exp Cell Res. 2019;385(2):111691. doi:10.1016/j.yexcr.2019.111691(IF:3.329)
[33] Yang B, Wang Y, Qin Q, et al. Polychlorinated Biphenyl Quinone Promotes Macrophage-Derived Foam Cell Formation. Chem Res Toxicol. 2019;32(12):2422-2432. doi:10.1021/acs.chemrestox.9b00184(IF:3.274)
[34] Yang B, Qin Q, Xu L, et al. Polychlorinated Biphenyl Quinone Promotes Atherosclerosis through Lipid Accumulation and Endoplasmic Reticulum Stress via CD36. Chem Res Toxicol. 2020;33(6):1497-1507. doi:10.1021/acs.chemrestox.0c00123(IF:3.184)
[35] Pan Z, Niu Y, Liang Y, Zhang X, Dong M. β-Ecdysterone Protects SH-SY5Y Cells Against 6-Hydroxydopamine-Induced Apoptosis via Mitochondria-Dependent Mechanism: Involvement of p38(MAPK)-p53 Signaling Pathway. Neurotox Res. 2016;30(3):453-466. doi:10.1007/s12640-016-9631-7(IF:3.140)
[36] Zhou Y, Huang N, Li Y, Ba Z, Zhou Y, Luo Y. Icaritin protects SH-SY5Y cells transfected with TDP-43 by alleviating mitochondrial damage and oxidative stress. PeerJ. 2021;9:e11978. Published 2021 Aug 10. doi:10.7717/peerj.11978(IF:2.984)
[37] Hu T, Li P, Luo Z, et al. Chloroquine inhibits hepatocellular carcinoma cell growth in vitro and in vivo. Oncol Rep. 2016;35(1):43-49. doi:10.3892/or.2015.4380(IF:2.301)
[38] An X, Ji B, Sun D. TRIM34 localizes to the mitochondria and mediates apoptosis through the mitochondrial pathway in HEK293T cells. Heliyon. 2020;6(1):e03115. Published 2020 Jan 9. doi:10.1016/j.heliyon.2019.e03115(IF:0.000)
[39] Huang T, Wang G, Hu Y, et al. Structural and functional abnormalities of penile cavernous endothelial cells result in erectile dysfunction at experimental autoimmune prostatitis rat. J Inflamm (Lond). 2019;16:20. Published 2019 Jul 25. doi:10.1186/s12950-019-0224-0(IF:0.000)
