生物发光细菌成像<em在体内</em
Summary
本文介绍了管理<em>勒克斯标签</em细菌对小鼠和随后的<em在体内</em分析IVIS生物发光成像。
Abstract
本视频介绍了使用全身生物发光成像（BLI）的细菌贩运活体小鼠为研究对象，利用细菌基因和细胞治疗癌症的重点。存在的细菌有吸引力的载体类用于癌症治疗，全身用药后，肿瘤内优先发展拥有一种天生的能力。细菌的设计，以表达勒克斯基因盒许可证BLI检测的细菌和兼肿瘤部位。肿瘤内随着时间的推移的位置和水平的细菌，可以容易地检查，在两维或三维可视化。的方法，是适用于范围广泛的细菌物种和肿瘤异种移植类型。本文介绍了发光细菌在皮下荷瘤小鼠的协议分析。可视化的共生细菌的胃肠道（GIT）中由BLI也被描述。这种功能强大，而且价格便宜，实时成像战略的代表中TS细菌在体内的背景下，特别是基因治疗，癌症研究，传染病研究的一个理想的方法。此视频概述了研究勒克斯标记E.程序大肠杆菌活体小鼠，展示的空间和时间的读出可以实现利用BLI的IVIS系统。
Protocol
1。诱发肿瘤对于常规的肿瘤诱导，悬浮在200μl的无血清培养基中的细胞的的最低致瘤剂量皮下注射（SC）的到感染的侧翼免费的6-8周龄的雌性Balb / C或无胸腺MF1-nu/nu小鼠的中N = 6（哈伦，英国牛津）（1×10 6 4T1细胞），使用21号注射器针头。用于接种的细胞的存活率大于95％，使用血细胞计数器和台盼蓝染料排除（Gibco）中通过目视计数来确定。 肿瘤成立后，肿瘤被允许的成?…
Discussion
在基因治疗的情况下，交付的基因治疗给患者使用生物制剂表现出极大的承诺3-5。如病毒，细菌先天的生物学性质允许高效的DNA传递到细胞或组织，特别是在癌症的上下文中。它已被证明细菌是自然能归巢到肿瘤全身给药时，所得的复制水平高的地方，无论是外部的（非侵入性的物种）或肿瘤细胞（病原体）内。癌特异性的细菌复制最初归因于缺氧的固体肿瘤的性质（低O 2水平）…
Divulgaciones
The authors have nothing to disclose.
Acknowledgements
作者要感谢支持本稿件相关的力士标记E.欧洲委员会第七框架计划（PIOF-GA-2009-255466）和爱尔兰健康研究委员会（HRA_POR/2010/138）。 大肠杆菌是一种礼物科马克·加恩，科克大学博士。
Materials
| Name of the reagent | Company | Catalogue number | Comments |
| 4T1 cell line | ATCC | CRL-2539 | Syngeneic breast cancer model derived from a spontaneously arising BALB/c mammary tumor |
| DMEM | Sigma-Aldrich | D6429 | Dulbecco’s Modified Eagle’s Medium |
| PBS | Sigma-Aldrich | D8537 | Phosphate Buffered Saline |
| Xenogen IVIS | Caliper Life Sciences | IVIS 100 for 2D imaging; IVIS Spectrum for 3D. | |
| Luria Broth Miller (LB) | Sigma-Aldrich | L2542 | Growth medium for E. coli |
| Erythromycin | Sigma-Aldrich | E5389 | Antibiotic |
| Streptomycin | Sigma-Aldrich | S9137 | Antibiotic |
| MF1nu/nu mice | Harlan (UK) | 069(nu)/070(nu/+) | Hsd:Athymic Nude-Foxn1nu |
| Balb/c mice | Harlan (UK) | 066 | Haplotype: H-2d |
| Gavage needle | Vet-tech Solutions (UK) | DE009 | 22G x 38mm straight gavage needle |
| Syringe for IV injection | BD BioSciences | 309309 – 1 ml | Insulin syringe with 28 G x ½ inch micro-fine IV needle. |
| Syringe for tumor inoculation | Braun | 9161376V | Omnifix 26 G x ½ inch needle |
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Tags
Bioluminescent Bacterial ImagingIn VivoWhole Body ImagingBLIBacterial TraffickingGene TherapyCell TherapyCancerLux Gene CassetteTumor DetectionTwo Or Three DimensionsBacterial SpeciesTumor XenograftSubcutaneous Tumor Bearing MiceVisualizationCommensal BacteriaGastrointestinal Tract (GIT)Real-time ImagingGene Therapy ResearchInfectious Disease ResearchLux-tagged E. ColiSpatial And Temporal ReadoutIVIS System
Citar este artículo
Baban, C. K., Cronin, M., Akin, A. R., O’Brien, A., Gao, X., Tabirca, S., Francis, K. P., Tangney, M. Bioluminescent Bacterial Imaging In Vivo. J. Vis. Exp. (69), e4318, doi:10.3791/4318 (2012).