Establishment of Patient-Derived Xenografts in Mice

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Patient-derived xenograft (PDX) models for cancer research have recently attracted considerable attention in both the academy and industry (Hidalgo et al., 2014; Wilding and Bodmer, 2014). PDX models have been developed from different tumor types including lung cancer to improve the drug development process. These models are used for pre-clinical drug evaluation and can be used for the predictive results of clinical outcomes because they conserve original tumor characteristics such as heterogeneity, complexity and molecular diversity (Kopetz et al., 2012). Additionally, PDX model provides the potential tool for the personalized drug therapy. In this protocol, we present methods for the establishment of PDX in mice using primary tumor tissues from patients with small cell lung cancer (SCLC).

Materials and Reagents

  1. Sterile alcohol prep pads (Covidien, catalog number: 6818 )
  2. Petri dishes (Corning, Falcon®, catalog number: 353003 )
  3. Tissue adhesive (3M, catalog number: 1469SB )
  4. 6 weeks old SCID mice (Charles River Laboratories International, catalog number: 236 ) or athymic nude mice (Envigo, catalog number: 069 )
  5. Ketamine hydrochloride/xylazine hydrochloride solution (Sigma-Aldrich, catalog number: K113 )
  6. Phosphate-buffered saline (PBS) (Mediatech, catalog number: 21-040-CV )
  7. 70% ethanol (Decon Labs, catalog number: 2401 )


  1. Stainless steel sterile scalpels (Integra LifeSciences, Miltex®, catalog number: 4-423 )
  2. Operating scissors (Sklar Surgical Instruments, catalog number: 13-1045 )
  3. Micro dissecting forceps (Roboz Surgical Instrument, catalog number: RS-5135 )
  4. Biological safety cabinet (Labconco, catalog number: 3460001 )
  5. CO2 chamber in animal facility
  6. Heating pad (Sunbeam Products, catalog number: 000765-500-000 )


  1. Collect pieces of primary tumors by surgery or biopsy procedures (F0).
  2. Anesthetize the SCID mice using ketamine hydrochloride/xylazine hydrochloride solution.
  3. Remove hair from the dorsal region.
  4. Cut the tumor (F0) into pieces.
  5. Make a shallow incision in the dorsal region of SCID mouse using a scissor and then implant subcutaneously two pieces of primary tumor into the interscapular fat pad of SCID mouse within 3 h after collecting primary tumor tissues from a patient. These mice in the engraftment phase are called F1 mice (SCID mice are used for F1 mice due to higher engraftment rates than those of athymic nude mice).
  6. Move the F1 mice to a warm area on heating pad and monitor them during recovery.
  7. Return F1 mice to the routine mouse sterile housing.
  8. When the tumor grows and reaches around 1.5 cm (4-10 weeks), prepare the additional transplant.
  9. Place the F1 mouse in the CO2 chamber for 5 min to euthanize it.
  10. Move the euthanized mouse into biological safety cabinet and clean the whole body of mouse using sterile alcohol prep pads.
  11. Cut off skin around the tumor area.
  12. Resect the tumor and transfer the tumor to a sterile Petri dish containing PBS (Video 1).
  13. Cut the tumor into even size piece (i.e., 2 mm3) (Video 1).

    Video 1. The cutting of tumor into small even piece for engraftment

    Figure 1. Establishment of PDX model. A. Photographs of representative small cell lung cancer (SCLC) PDX in mice (F3 mice). B. H&E histology of tumor tissues from SCLC PDX in mice. TKO stands for Dr. Taofeek K. Owonikoko who provided primary lung tumor tissues.

  14. Anesthetize the athymic nude mice like described in step 2.
  15. Implant a piece of tumor into each experimental athymic nude mouse like described in step 5. Apply tissue adhesive to cut surfaces of the skin. Mice in this expansion phase are called F2 mice. After 2-4 weeks, the tumors could be visible as in Figure 1 and Video 2.
  16. Move the F2 mice to a warm area and monitor them like described step 5.
  17. Return the F2 mice to the routine mouse sterile housing.
  18. When the tumor volume reaches 100-150 mm3, mice can be used for downstream application.

    Video 2. The tumor implantation in mice


  1. Ensure that all procedures should be undertaken in a biological safety cabinet.
  2. Ensure that any necrotic tumor tissue is not used for transplant (Figure 2).
  3. Ensure that only early-passage PDXs (less than the 5th passage) can be used for the studies.

    Figure 2. Photographs of representative non-necrotic tissue and necrotic tissue


We thank Dr. Taofeek K. Owonikoko, Assistant Professor and Guojing Zhang, Research Specialist at Emory University for providing primary lung tumor tissues. This work is supported by NCI, National Institutes of Health grants 1R01CA193828, 2R01CA136534 and 1R01CA200905-01A1.


  1. Hidalgo, M., Amant, F., Biankin, A. V., Budinska, E., Byrne, A. T., Caldas, C., Clarke, R. B., de Jong, S., Jonkers, J., Maelandsmo, G. M., Roman-Roman, S., Seoane, J., Trusolino, L. and Villanueva, A. (2014). Patient-derived xenograft models: an emerging platform for translational cancer research. Cancer Discov 4(9): 998-1013.
  2. Kopetz, S., Lemos, R. and Powis, G. (2012). The promise of patient-derived xenografts: the best laid plans of mice and men. Clin Cancer Res 18(19): 5160-5162.
  3. Wilding, J. L. and Bodmer, W. F. (2014). Cancer cell lines for drug discovery and development. Cancer Res 74(9): 2377-2384.


用于癌症研究的患者衍生的异种移植(PDX)模型最近在学术和工业中引起了相当大的关注(Hidalgo等人,2014; Wilding和Bodmer,2014)。 PDX模型已经从不同的肿瘤类型开发,包括肺癌,以改善药物开发过程。 这些模型用于临床前药物评价,并且可用于临床结果的预测结果,因为它们保留原始肿瘤特征,例如异质性,复杂性和分子多样性(Kopetz等人,2012) 。 此外,PDX模型为个性化药物治疗提供了潜在的工具。 在该协议中,我们提出了使用来自小细胞肺癌(SCLC)患者的原发性肿瘤组织在小鼠中建立PDX的方法。


  1. 无菌酒精制备垫(Covidien,目录号:6818)
  2. 培养皿(Corning,Falcon ,目录号:353003)
  3. 组织粘合剂(3M,目录号:1469SB)
  4. 6周龄SCID小鼠(Charles River Laboratories International,目录号:236)或无胸腺裸鼠(Envigo,目录号:069)
  5. 盐酸氯胺酮/盐酸赛拉嗪溶液(Sigma-Aldrich,目录号:K113)
  6. 磷酸盐缓冲盐水(PBS)(Mediatech,目录号:21-040-CV)
  7. 70%乙醇(Decon Labs,目录号:2401)


  1. 不锈钢无菌手术刀(Integra LifeSciences,Miltex ,目录号:4-423)
  2. 操作剪刀(Sklar Surgical Instruments,目录号:13-1045)
  3. 微解剖钳(Roboz Surgical Instrument,目录号:RS-5135)
  4. 生物安全柜(Labconco,目录号:3460001)
  5. 动物设施中的CO 2腔室
  6. 加热垫(Sunbeam Products,目录号:000765-500-000)


  1. 通过手术或活检程序(F0)收集原发性肿瘤。
  2. 麻醉SCID小鼠使用氯胺酮盐酸盐/盐酸赛拉嗪溶液
  3. 从背部区域取出头发。
  4. 切割肿瘤(F0)成块。
  5. 使用剪刀在SCID小鼠的背部区域做一个浅切口,然后在从患者收集原发性肿瘤组织后3小时内将两片原发性肿瘤皮下植入SCID小鼠的肩胛间脂肪垫。这些处于移植阶段的小鼠称为F1小鼠(SCID小鼠用于F1小鼠,因为其移植率高于无胸腺裸鼠)。
  6. 将F1小鼠移动到加热垫上的温暖区域,并在恢复期间监测它们
  7. 将F1小鼠返回常规小鼠无菌室
  8. 当肿瘤生长并达到约1.5cm(4-10周)时,准备额外的移植
  9. 将F1小鼠放在CO 2室中5分钟以使其安乐死。
  10. 将安乐死的小鼠移入生物安全柜,并使用无菌酒精制备垫清洁整个小鼠
  11. 切除肿瘤区域周围的皮肤。
  12. 切除肿瘤并将肿瘤转移到含有PBS的无菌培养皿(视频1)
  13. 将肿瘤切成均匀尺寸(,<2mm> 3 )(视频1)。

    图1. PDX模型的建立 A.小鼠(F3小鼠)中代表性小细胞肺癌(SCLC)PDX的照片。 B.来自SCLC PDX的肿瘤组织在小鼠中的H& E组织学。 TKO代表提供原发性肺肿瘤组织的Taofeek K. Owonikoko博士
  14. 麻醉无胸腺的裸鼠,如步骤2所述。
  15. 如步骤5中所述,将一块肿瘤植入每个实验无胸腺裸鼠。将组织粘合剂应用于皮肤的切割表面。在该扩增期的小鼠称为F2小鼠。 2-4周后,肿瘤可见,如图1和视频2
  16. 将F2鼠标移动到温暖的区域,按照步骤5所述进行监控。
  17. 将F2小鼠返回常规小鼠无菌室。
  18. 当肿瘤体积达到100-150mm 3 时,小鼠可用于下游应用


  1. 确保所有程序都应在生物安全柜中进行。
  2. 确保任何坏死肿瘤组织不用于移植(图2)。
  3. 确保只有早期传代的PDX(小于5 th 段)可用于研究。



我们感谢埃默里大学研究专家助理教授Taofeek K. Owonikoko博士和提供原发性肺肿瘤组织的郭国景博士。这项工作由NCI,国家卫生研究院拨款1R01CA193828,2R01CA136534和1R01CA200905-01A1支持。


  1. Hidalgo,M.,Amant,F.,Biankin,AV,Budinska,E.,Byrne,AT,Caldas,C.,Clarke,RB,de Jong,S.,Jonkers,J.,Maelandsmo,GM,Roman-Roman ,S.,Seoane,J.,Trusolino,L.和Villanueva,A。(2014)。  患者衍生的异种移植物模型:用于翻译癌症研究的新兴平台 Cancer Discov 4(9):998-1013。
  2. Kopetz,S.,Lemos,R。和Powis,G。(2012)。  患者衍生的异种移植的希望:小鼠和男人最好的计划。 Clin Cancer Res 18(19):5160-5162。
  3. Wilding,JL和Bodmer,WF(2014)。  癌症用于药物发现和开发的细胞系。 Cancer Res 74(9):2377-2384。
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Copyright: © 2016 The Authors; exclusive licensee Bio-protocol LLC.
引用:Park, D., Wang, D., Chen, G. and Deng, X. (2016). Establishment of Patient-Derived Xenografts in Mice. Bio-protocol 6(22): e2008. DOI: 10.21769/BioProtoc.2008.

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