描述
CYTO-ID®自噬检测试剂盒
产品信息
品牌 |
产品名称 | 产品编号 | 规模 | 价格（元） |
| Enzo Life | CYTO-ID® Autophagy detection kit CYTO-ID®自噬检测试剂盒 | ENZ-51031-K200 | 200 tests | 6226.25 |
| Enzo Life | CYTO-ID® Autophagy detection kit CYTO-ID®自噬检测试剂盒 | ENZ-51031-0050 | 50 tests | 3220.5 |
品特点：
无需转染
- 专有染料包括专用于选择性染色自噬囊泡的可滴定部分
- 用已知的自噬活性抑制剂和激活剂验证的方案
- 快速量化天然异质细胞群中的自噬
- 无需进行LC3-GFP转染所需的费时费力的转染效率验证
- 选择性和全面染色，允许测量和区分自噬通量和自噬溶酶体积累
- 溶酶体染色可忽略不计，减少了其他染料所见的背景
- 促进自噬激活剂和抑制剂的高通量筛选
产品详情
| 应用： | 流式细胞术、荧光显微镜、荧光检测、HTS | |
| 应用说明： | CYTO-ID ®自噬检测试剂盒为通过荧光显微镜、流式细胞术和酶标仪监测活细胞中的自噬提供了一种快速、特异性和定量的方法。 | |
| 质量控制： |
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| 数量： | 对于 -K200 大小： 200 次流式细胞术检测、250 次显微镜检测或 3 x 96 孔微孔板检测。对于 -0050 尺寸： 50 次流式细胞术检测、60 次显微镜检测或 1 x 96 孔微孔板检测。 |
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| 使用/稳定性： | 在妥善储存的情况下，试剂盒组件自收到之日起可稳定保存一年。 | |
| 处理： |
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| 船运： | 在蓝冰上发货 | |
| 短期储存： |
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| 内容： |
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| 科学背景： | 自噬是一种应激诱导的保护机制。在基础条件下活性较低，真核细胞通过溶酶体介导的细胞内容物大量降解，在遇到某些不利条件（例如营养耗尽和化学或环境压力）时利用该机制。自噬活性增加在癌症、神经变性、心血管疾病和糖尿病病理学中的作用已得到广泛认可和普遍研究。如果溶酶体功能被抑制，自噬流的诱导可以通过自噬囊泡的增强积累来可视化，阻止这些囊泡的去除。 | |
| 技术信息/产品说明： |
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| 协议： | 可在 bioprotocol.org 上找到原代 BMDC 中 FC 的详细协议：M. Stankov 等人 在原代细胞中使用 CYTO-ID 染色对自噬活性进行流式细胞分析。 |
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| 监管状态： | RUO – 仅供研究使用 |
产品参数：
CYTO-ID®自噬检测试剂盒使用选择性标记积累的自噬液泡的新型染料测量自噬液泡并监测溶酶体抑制的活细胞中的自噬通量。该染料通过鉴定可滴定的功能部分进行了优化，这些部分允许溶酶体的染色最少，同时在掺入前自噬体、自噬体和自噬溶酶体（自噬溶酶体）时表现出明亮的荧光。该测定提供了一种无需细胞转染即可监测活细胞自噬的快速定量方法。
作用机制：
该探针是一种阳离子两亲示踪剂 (CAT) 染料，它以与诱导磷脂质沉着症的药物类似的方式迅速分配到细胞中。仔细选择染料上的可滴定功能部分可防止其在溶酶体中积累，但可以标记与自噬途径相关的液泡。
自噬示意图。细胞质物质被不断膨胀的膜囊（吞噬细胞）隔离，从而形成双膜囊泡，即自噬体。自噬体的外膜随后与溶酶体融合，内部物质在自噬溶酶体中降解。图中还描述了各种自噬调节剂。
使用 CYTO-ID® 自噬检测试剂盒进行基于流式细胞术的自噬分析：未诱导的对照（红线峰）和 10uM 他莫昔芬 (ALX-550-095) 处理的（蓝色填充峰）Jurkat 细胞（T 细胞白血病）用过的。处理 18 小时后，将细胞加载 CYTO-ID® Green 检测试剂，然后通过流式细胞术进行分析，无需洗涤。结果由直方图叠加呈现。对照细胞也被染色，但大多显示低荧光。在用 10uM 他莫昔芬处理 18 小时的样品中，CYTO-ID® Green 染料信号增加了约 2 倍，表明他莫昔芬导致 Jurkat 细胞的自噬增加。
使用 CYTO-ID® 自噬检测试剂盒进行基于流式细胞术的自噬分析：未诱导的对照（红线峰）和 10uM 他莫昔芬 (ALX-550-095) 处理的（蓝色填充峰）Jurkat 细胞（T 细胞白血病）用过的。处理 18 小时后，将细胞加载 CYTO-ID®Green 检测试剂，然后通过流式细胞术进行分析，无需洗涤。结果由直方图叠加呈现。对照细胞也被染色，但大多显示低荧光。在用 10uM 他莫昔芬处理 18 小时的样品中，CYTO-ID®Green 染料信号增加了约 2 倍，表明他莫昔芬导致 Jurkat 细胞的自噬增加。
HepG2 细胞与雷帕霉素（一种 mTOR 激酶抑制剂）的过夜孵育会导致 CYTO-ID®染料信号增加。
消除非特异性溶酶体染色导致的背景。CYTO-ID®Green 染料消除了使用单丹糖基尸胺 (MDC)（下图）的其他基于溶酶体营养染料的检测所见的溶酶体背景染色。CYTO-ID®自噬试剂盒无需使用 350 nm 紫外激光进行活细胞分析，并且与 Hoechst 染料兼容，可在显微镜应用中进行共标记。
自噬积累和自噬通量的可视化。通过荧光显微镜观察，自噬液泡的积累和通量都可以通过 CYTO-ID® Autophagy Green 染料进行检测。HeLa 细胞用 0.2% DMSO (A) 模拟诱导或用 100 uM 盐酸可乐定 (B)、5 uM 盐酸洛哌丁胺 (C) 或 1 uM PP242 水合物 (D) 在 37°C 下诱导 12 小时。处理后，将细胞与 CYTO-ID® Green 检测试剂在 37°C下孵育10 分钟，然后用测定缓冲液洗涤。细胞核用 Hoechst 33342 染料复染成蓝色。
未转染的自噬曲线。图 1A：稳定表达 GFP-LC3 转染细胞系的 CHO 细胞导致对照与饥饿细胞群的基线分离相对较差，从而难以量化自噬。图改编自 Shvets E、Fass E、Elazar Z。 图 1B：CYTO-ID®自噬检测试剂盒专门标记独立于 LC3 蛋白的自噬泡，无需转染。HeLa 细胞经过饥饿和恢复，然后用 CYTO-ID®Green 检测试剂标记。该染料能够清晰检测和量化与自噬诱导直接相关的自噬和前自噬液泡。
无需转染即可对自噬泡进行省时、快速和全面的标记。为了展示 CYTO-ID® Green 检测试剂的优势，首先用 RFP-LC3 表达载体转染 HeLa 细胞，用 10 µM 他莫昔芬处理过夜，然后用 CYTO-ID ® Green 检测试剂染色。与基于过夜转染的检测不同，CYTO-ID® Green 检测试剂方法可在 15-30 分钟内标记 100% 的细胞。图 A：绿色信号，表明自噬囊泡的CYTO-ID®Green 染色；图 B：成功转染细胞亚群中的 RFP-LC3 表达（红色）；图 C：合成图像，显示 CYTO-ID®绿色染料标记的囊泡与自噬体的特定标记 LC3 共定位。
CYTO-ID®自噬检测试剂盒
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