Fluorescence lifetime imaging of activatable target specific molecular probes.
Fluorescence lifetime imaging of activatable target specific molecular probes.
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DOI：
10.1002/cmmi.360
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发表时间：
2010-01
影响因子：
--
通讯作者：
Kobayashi, Hisataka
中科院分区：
文献类型：
--
作者：
Alford, Raphael;Ogawa, Mikako;Hassan, Moinuddin;Gandjbakhche, Amir H.;Choyke, Peter L.;Kobayashi, Hisataka
利用荧光标记的自猝灭单克隆抗体进行体内光学成像，通过靶细胞内的结合和内化激活，可获得优异的靶背景比。我们假设这些分子探针可以利用荧光寿命成像（FLI）准确地报告细胞内化。合成两种成像探针，由抗体曲妥珠单抗（靶向HER2/neu）以1:8或1:1的比例偶联Alexa Fluor750组成。每个偶联物的荧光强度和寿命最初在内体ph值测定。由于1:8共轭物是自猝灭的，因此在暴露于已知的猝灭剂SDS后，还测定了每个探针的荧光寿命。采用3T3/HER2+和BALB/3T3 （HER2−）细胞系进行体外成像实验。荧光寿命的变化与温度和时间依赖性细胞内化相关。双侧腹肿瘤小鼠的体内成像研究[3T3/HER2+和BALB/3T3 (HER2−)]检测到FLI的微小差异。总之，荧光寿命成像可监测体外靶向活化抗体-荧光基团偶联物的内在化。挑战仍然是适应这种方法在体内成像。
In vivo optical imaging using fluorescently labeled self-quenched monoclonal antibodies, activated through binding and internalization within target cells, results in excellent target-to-background ratios. We hypothesized that these molecular probes could be utilized to accurately report on cellular internalization with fluorescence lifetime imaging (FLI). Two imaging probes were synthesized, consisting of the antibody trastuzumab (targeting HER2/neu) conjugated to Alexa Fluor750 in ratios of either 1:8 or 1:1. Fluorescence intensity and lifetime of each conjugate were initially determined at endosomal pHs. Since the 1:8 conjugate is self-quenched, the fluorescence lifetime of each probe was also determined after exposure to the known dequencher SDS. In vitro imaging experiments were performed using 3T3/HER2+ and BALB/3T3 (HER2−) cell lines. Changes in fluorescence lifetime correlated with temperature- and time-dependent cellular internalization. In vivo imaging studies in mice with dual flank tumors [3T3/HER2+ and BALB/3T3 (HER2−)] detected a minimal difference in FLI. In conclusion, fluorescence lifetime imaging monitors the internalization of target-specific activatable antibody–fluorophore conjugates in vitro. Challenges remain in adapting this methodology to in vivo imaging.
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影响因子：
5.7
作者：
Ogawa M;Regino CA;Choyke PL;Kobayashi H
通讯作者：
Kobayashi H
影响因子：
11.5
作者：
Barrett, Tristan;Koyama, Yoshinori;Kobayashi, Hisataka
通讯作者：
Kobayashi, Hisataka
影响因子：
2.4
作者：
Gannot, I;Ron, I;Gandjbakhche, A
通讯作者：
Gandjbakhche, A
影响因子：
4.7
作者：
Hama, Yukihiro;Urano, Yasuteru;Kobayashi, Hisataka
通讯作者：
Kobayashi, Hisataka
影响因子：
4.7
作者：
Ogawa, Mikako;Kosaka, Nobuyuki;Kobayashi, Hisataka
通讯作者：
Kobayashi, Hisataka