常见的流式细胞术基本原理及应用.ppt

* * 尿嘧啶核苷 * 磷酯酰丝氨酸 * * * * * * * * 使用 BD? Cytometric Bead Array(CBA) 系统你可以: ? 从单一小体积样本中得到多项检测结果 ? 对所有的检测项目，只需要制备一个标准混合物来绘制标准曲线 ? 避免人为造成的酶联放大的假阳性信号产生 ? 用更少的时间和精力，却得到大量的结果 ? 如果是用488nm和635nm两根激光器来实验，实验条件更容易建立 ? 使用配有HTS选件的BD流式细胞仪可以做到自动平板上样和高通量检测 目前CBA主要提供的检测内容 可溶性细胞因子 细胞凋亡相关蛋白 磷酸化蛋白 BD CBA Flex Set A B C D E F G H I 1 2 3 4 5 6 7 8 9 FL4-H or Red-A FL3-H or NIR-A Old Format BD CBA Beads BD CBA Flex Set Beads 更为灵活强大的CBA Flex Set 最多可同时检测72个项目 可自由选择搭配检测项目 检测项目范围更广泛 需要488nm和635nm两种波长的激光器 A B C D E F G H I 1 2 3 4 5 6 7 8 9 FL4-H or Red-A FL3-H or NIR-A 9个指标同时检测活化T细胞 1 2 3 4 5 6 7 8 9 1. Itk (Y511) 2. ERK (T202/Y204) 3. JNK (T183/Y185) 4. P38 (T180/Y182) 5. PLCg (Y783) 6. ZAP70 (Y319) 7. LAT (Y171) 8. c-Jun (S63) 9. RSK (S380) Kinetics of Jurkat Cell Activation With Anti-CD3/CD28 1 10 100 FOLD INCREASE IN UNITS/ML 0 5 10 15 20 TIME (MINUTES) RSK (S380) Jun (S63) LAT (Y171) Itk (Y511) ZAP-70 (Y319) PLCg (Y783) P38 (T180/Y182) JNK (T183/Y185) ERK (T202/Y204) P-Specific Antibodies Monitoring the T Cell Activation Pathway by CBA (Control) Lck ZAP-70 P Itk P P MAPKKK P IKK P NF-kB Ras Grb2 S O S Rac P P PLC P P DAG PKC P V A V SLP-76 P LAT P P P P P P P P P P P P Ras MEK ERK JNK p38 MKK4,7 MKK3,6 MEKK1-4 Elk Jun ATF2 MEKK4 LAT P P RSK Cells were activated with anti-CD3/CD28 for 2 minutes. SDS was added to a final concentration of 1% and the material was placed in a boiling water bath for 5 minutes. Monitoring the T Cell Activation Pathway by CBA (PD-98059) Jurkat cells were pre-incubated with 200 mM PD-98059 (MEK inhibitor) for 20 minutes before being activated activated with anti-CD3/CD28 for 2 minutes. SDS was added to a final concentration of 1% and the material was placed in a boiling water bath for 5 minutes. Monitoring the T Cell Activation Pathway by CBA (PP2) Jurkat cells were pre-incubated with 10 mM PP2 (Src family kinase inhibitor) for 20 minutes before being activated activated with anti-CD3/CD28 for 2 minutes. SDS was added to a final concentrati