Detection and measurement of in vitro gene transfer by gamma camera imaging.
Detection and measurement of in vitro gene transfer by gamma camera imaging.
复制标题
通过伽玛相机成像检测和测量体外基因转移。
DOI：
10.1038/sj.gt.3301391
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发表时间：
2001
期刊：
影响因子：
5.1
通讯作者：
Rogers,BE
中科院分区：
文献类型：
--
作者：
Zinn,KR;Chaudhuri,TR;Buchsbaum,DJ;Mountz,JM;Rogers,BE
这项工作的目的是开发一种高容量方法，对细胞培养板中单层生长的癌细胞的基因转移进行成像。用于体外检测基因转移的灵敏且高容量的核成像方法将允许在不同条件下的不同细胞系中快速验证载体。人类癌细胞系（A-427 非小细胞肺癌、SKOV3.ip1 卵巢、MDA-MB-468 乳腺癌和 BxPC-3 胰腺）被编码人 2 型生长抑素受体 (Ad-hSSTr2) 的复制无能腺病毒载体感染。通过对内化的 99m Tc 标记的 hSSTr2 结合肽（P2045，Diatide，Inc.）进行成像来检测细胞中 hSSTr2 报告蛋白的表达。成像提供了内部结合 99m Tc 的准确测量，成像感兴趣区域 (ROI) 分析和伽玛计数器测量结果的等效性证明了这一点。内部结合的 99m Tc-P2045 与基因转移后 hSSTr2 阳性细胞的百分比线性相关 (R 2 = 0.98)。过量的 P2045 阻断了 99m Tc-P2045 的结合和内化，表明该技术的特异性。最多可以同时对四个 96 孔板进行成像，从而证明了系统的高容量。随着新载体的开发，这种新颖的体外方法提供了一种测试增强基因转移的新方法。
The purpose of this work was to develop a high capacity method to image gene transfer to cancer cells growing as monolayers in cell culture plates. A sensitive and high capacity nuclear-imaging method for detection of gene transfer in vitro will allow rapid validation of vectors in different cell lines under various conditions. Human cancer cell lines (A-427 non-small cell lung, SKOV3. ip1 ovarian, MDA-MB-468 breast, and BxPC-3 pancreatic) were infected with a replication-incompetent adenoviral vector encoding the human type 2 somatostatin receptor (Ad-hSSTr2). Expression of the hSSTr2 reporter protein in cells was detected by imaging an internalized 99m Tc-labeled, hSSTr2 binding peptide (P2045, Diatide, Inc.). Imaging provided an accurate measure of internally bound 99m Tc as evidenced by equivalence of results for imaging region of interest (ROI) analyses and gamma counter measurements. Internally bound 99m Tc-P2045 was linearly correlated (R 2= 0.98) with the percentage of hSSTr2-positive cells following gene transfer. Excess P2045 blocked binding and internalization of the 99m Tc-P2045, indicating the specificity of the technique. Up to four 96-well plates could be imaged simultaneously, thereby demonstrating the high capacity of the system. This novel in vitro approach provides a new method to test enhanced gene transfer as new vectors are developed.