Summary

生物发光细菌成像<em在体内</em

Published: November 04, 2012
doi:

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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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).

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