Claudio R. Thoma, Alexandre Matov, Katrin L. Gutbrodt, Christian R. Hoerner, Zlatko Smole, Wilhelm Krek, Gaudenz Danuser (2011) CIL:31263, Homo sapiens, epithelial cell, renal cell carcinoma. CIL. Dataset

Immunofluorescence images used to investigate stabilization of microtubules (MTs) by Von Hippel-Lindau tumor suppressor protein (pVHL). VHL-deficient RCC-4 cells with empty vector have a lower alpha-tubulin (MTs; green) signal relative to GSK-3beta (red; an internal fluorescence control), compared with RCC-4 cells expressing pVHL30, suggesting that VHL stabilizes MTs. RCC-4 cells were infected with a retroviral vector to express pVHL30, fixed in 3% paraformaledehyde and permeabilized in 0.2% Triton X-100. Primary antibodies used were: monoclonal anti-GSK3beta and polyclonal anti-alpha tubulin. Secondary antibodies used were: anti-rabbit FITC and anti-mouse Texas-Red. Stacks of 5 z planes were acquired with a Leica SP2 system using the 63x 1.4NA, Oil, HCX Plan-Apo Objective and the distance between z planes is 1 micron. This stack corresponds to J Cell Biol. 2010. 190: 991-1003--Fig 1C (left panels) and the images of RCC-4 expressing pVHL30 are in Fig 1C (right panels) and CIL# 31263.

本研究采用了免疫荧光图像以探究冯·希佩尔-林道肿瘤抑制蛋白（pVHL）对微管（MTs）稳定性的影响。与携带空载体的VHL缺陷型RCC-4细胞相比，表达pVHL30的RCC-4细胞在α-微管蛋白（MTs；绿色荧光信号）相对于GSK-3beta（红色荧光信号，作为内源荧光对照）的信号强度更低，这一发现暗示了VHL能够稳定微管结构。RCC-4细胞通过逆转录病毒载体感染以表达pVHL30，随后使用3%多聚甲醛固定，并以0.2%的Triton X-100进行渗透处理。本研究中使用的初级抗体包括抗GSK3beta的单克隆抗体和抗α-微管蛋白的多克隆抗体，而二级抗体则包括抗兔FITC和抗鼠Texas-Red。利用Leica SP2系统，配备63x 1.4NA油镜HCX Plan-Apo物镜，采集了5层z平面的图像堆栈，层间距为1微米。该图像堆栈对应于《细胞》杂志2010年第190期第991-1003页的图1C（左侧面板）以及表达pVHL30的RCC-4细胞的图像，见图1C（右侧面板）和CIL# 31263。