T细胞亚群检测资料.pptVIP

* * 流式细胞仪通过前向散射与侧向散射、荧光信号这两种属性将细胞进行分群、分析的。 * Dichroic,二色的 fliters Bandpass filters * Their fluorescence emission wavelengths are: FITC 530 nm, PE 575 nm, PE–CY5 670 nm PE–TR 613 nm, phycoerythrin–cyanine 5 tandem complex:PE-CY5 fluorescein isothiocyanate FITC , phycoerythrin PE , * Their fluorescence emission wavelengths are: FITC 530 nm, PE 575 nm, PE–CY5 670 nm PE–TR 613 nm, phycoerythrin–cyanine 花青 5 tandem complex:PE-CY5 fluorescein isothiocyanate FITC ,异硫氰酸荧光素 phycoerythrin PE ,藻红蛋白 Detection of T cell subtypes by Flow Cytometry * BD公司流式细胞仪 FACSCalibur LSRII FACSCanto FACSCount FACSVantageDiVa FACSAria * What is Flow Cytometry? Principals and Applications FLOW CYTOMETRY FLOW FLUID CYTO CELLS METRY MEASUREMENT Definition Flow Cytometry * Measuring properties of cells in flow Flow sorting * Sorting separating cells based on properties measuring in flow * Also called Fluorescence-Activated Cell Sorting FACS * FSC and SSC Forward Scatter FSC ---Approximate cell size Side Orthogonal Scatter SSC --- Cell complexity or granularity Scatter are used for preliminary identification of cells. In a peripheral blood sample, lymphocyte, monocyte and granulocyte populations can be defined on the basis of forward and side scatter. Forward and side scatter are used to exclude debris and dead cells. * FSC与SSC检测器 * FSC and SSC * FSC and SSC are used for preliminary identification of cells. * CD8 Step 1 Direct immunofluorescent Staining Step 2 Flow cytometry analysis Content Materials Mouse spleen Eppendorf tubes PBS Red blood cell lysing solution Anti-mouse CD3/FITC Anti-mouse CD4/PE Anti-mouse CD8/APC Flow cytometry machine FACSCalibur/ LSRII * Fluorescence emission wavelengths fluorescein isothiocyanate FITC , 530 nm , phycoerythrin PE , 575 nm phycoerythrin–cyanine 5 tandem complex:PE-CY5 670 nm * 流式细胞术检测小鼠脾脏T细胞亚群 * 基本步骤 小鼠脾细胞的获得 裂解红细胞、并细胞计数 荧光抗体染色 流式细胞术检测T细胞 * 小鼠脾脏T细胞亚群的检测（1） 颈椎脱臼处死小鼠，放入70% 的酒精浸泡消毒1min。