BD流式全面精选.ppt

* Some fluorescent proteins from Clonetech, our sister division. In this case 4 different cell-lines expression 4 different kinds of fluorescent proteins. DS red and Zsyellow of the blue laser (although zsyellow emits right between FITC and PE and is not that useful). But also look at the HCRed, emitting only in the red laser, and AmCyan in the violet laser, with just a little overlap in FITC. These pictures are uncompensated by the way. Optical Filter for FP on Aria ? ran on the FACSAria at 70psi. Standard configuration: ? DsRed: 488 laser, 585/42 filter ? ZsYellow: 488 laser, 530/30 filter (also shows in the 585/42) ? HcRed: 633 laser, 660/20 filter ? AmCyan: 407 laser, 450/40? (or 530/30, Im a little confused about these two, but you can check the emission spectrum and see which one would have made more sense) FCM in Molecular Biology 流式检测分子表型 特异的核酸序列 SNP 端粒长度 荧光报告蛋白检测基因表达情况 Fluorescent Proteins DsRed ZsYellow HcRed AmCyan 端粒长度的检测 端粒的长度和端粒酶的活性是现在研究的重点。因为它们被发现和细胞复制以及一些疾病的变化相关。 可以通过FISH，用带有荧光的核酸探针标记上端粒序列，通过流式细胞仪检测端粒的长度 流式分选应用举例 自然杀伤细胞分选 基因转染细胞的分选 绿色萤光蛋白分析（GFP）：左图显示Hela细胞转染了减毒Mengo病毒vM16，其中含转染后8小时与病毒L肽链融合的GFP。直方图显示59％细胞表达该肽链，并显示不同的表达水平。重叠的红线表示阴性对照。 * The answer is A. * This is a dot plot of Red vs. Far Red data for cells that have been stained with antibodies conjugated to fluorochromes. In this dot plot, APC is conjugated to Antbody A, while PE-Cy7 is conjugated to Antibody B. In this case, the fluorescence intensity is indicative of the antigen A and B expression in the cells. The more to the right of the scale the particle appears, the higher its fluorescence intensity. By convention, the lower left quadrant displays cells that are negative for both of these antigens. Explain the other quadrants. Which of the three populations has the most Antibody A binding sites? Answer: Lower Right. * The sample stream is injected into the center of the sheath steam in the optical cuvette and the particles are carried upward in a single file past the laser