Nanobody-coupled microbubbles as novel molecular tracer.
Nanobody-coupled microbubbles as novel molecular tracer.
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DOI：
10.1016/j.jconrel.2011.12.007
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发表时间：
2012-03-10
期刊：
影响因子：
--
通讯作者：
Devoogdt N
中科院分区：
文献类型：
--
作者：
Hernot S;Unnikrishnan S;Du Z;Shevchenko T;Cosyns B;Broisat A;Toczek J;Caveliers V;Muyldermans S;Lahoutte T;Klibanov AL;Devoogdt N
骆驼衍生的单结构域抗体片段（~ 15 kDa），称为纳米抗体，是一类新的分子示踪剂，其通常以纳摩尔亲和力对其靶点进行鉴定，并且易于定制用于分子成像和药物递送应用。我们假设它们非常适合靶向微泡（μ B）的设计，旨在开发和表征eGFP和VCAM-1靶向μ B。抗eGFP（cAbGFP 4）和抗VCAM-1（cAbVCAM 1 -5）纳米抗体在细菌中被位点特异性生物素化。这种代谢生物素化方法产生功能性纳米抗体，其中一个生物素位于分子的抗原结合区的远端位点。生物素化的纳米抗体通过链霉亲和素-生物素桥接与生物素化的脂质μ B偶联。通过荧光显微镜检测μ B-cAbGFP 4识别eGFP的能力作为原理验证，并证实eGFP与μ B-cAbGFP 4的特异性结合。动态流动室研究证明了μ B-cAbVCAM 1 -5在快速流动（高达5达因/cm 2）中结合VCAM-1的能力。在MC 38荷瘤小鼠（n=4）中进行体内靶向研究。静脉内注射μ B-cAbVCAM 1 -5或对照μ B-cAbGFP 4，并使用造影剂特异性超声成像模式进行成像。在注射后10分钟测量肿瘤中的回声强度。与对照μ B相比，μ B-cAbVCAM 1 -5显示增强的信号（p<0.05）。利用纳米抗体的代谢和位点特异性生物素化，描述了开发纳米抗体偶联的μBs的方法。VCAM-1靶向μBs作为新型分子超声造影剂的应用在体外和体内都得到了证实。
Camelid-derived single-domain antibody-fragments (~15kDa), called nanobodies, are a new class of molecular tracers that are routinely identified with nanomolar affinity for their target and that are easily tailored for molecular imaging and drug delivery applications. We hypothesized that they are well-suited for the design of targeted microbubbles (μBs) and aimed to develop and characterize eGFP- and VCAM-1-targeted μBs. Anti-eGFP (cAbGFP4) and anti-VCAM-1 (cAbVCAM1-5) nanobodies were site-specifically biotinylated in bacteria. This metabolic biotinylation method yielded functional nanobodies with one biotin located at a distant site of the antigen-binding region of the molecule. The biotinylated nanobodies were coupled to biotinylated lipid μBs via streptavidin-biotin bridging. The ability of μB-cAbGFP4 to recognize eGFP was tested as proof-of-principle by fluorescent microscopy and confirmed the specific binding of eGFP to μB-cAbGFP4. Dynamic flow chamber studies demonstrated the ability of μB-cAbVCAM1-5 to bind VCAM-1 in fast flow (up to 5 dynes/cm2). In vivo targeting studies were performed in MC38 tumor-bearing mice (n=4). μB-cAbVCAM1-5 or control μB-cAbGFP4 were injected intravenously and imaged using a contrast-specific ultrasound imaging mode. The echo intensity in the tumor was measured 10 minutes post-injection. μB-cAbVCAM1-5 showed an enhanced signal compared to control μBs (p<0.05). Using metabolic and site-specific biotinylation of nanobodies, a method to develop nanobody-coupled μBs was described. The application of VCAM-1-targeted μBs as novel molecular ultrasound contrast agent was demonstrated both in vitro and in vivo.
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影响因子：
6.7
作者：
Anderson CR;Rychak JJ;Backer M;Backer J;Ley K;Klibanov AL
通讯作者：
Klibanov AL
DOI：
10.1016/s1050-3862(99)00046-7
发表时间：
1999-12-31
期刊：
BIOMOLECULAR ENGINEERING
影响因子：
--
作者：
Chapman-Smith, A;Cronan, JE
通讯作者：
Cronan, JE
影响因子：
37.8
作者：
Ellegala, DB;Poi, HL;Lindner, JR
通讯作者：
Lindner, JR
影响因子：
10.8
作者：
Geers, Bart;Lentacker, Ine;De Smedt, Stefaan C.
通讯作者：
De Smedt, Stefaan C.
影响因子：
8
作者：
Dumoulin, M;Conrath, K;Matagne, A
通讯作者：
Matagne, A