This work is supported by grants from (1) the National Nature Science Foundation (81973877 and 82174408); (2) Shanghai Top Priority Research Center construction project (2022ZZ01009); (3) National Key R&#38;D Program of China (2020YFE0201600); (4) Shanghai Collaborative Innovation Center of Industrial Transformation of Hospital TCM Preparation and (5) Research Projects within Budget of Shanghai University of Traditional Chinese Medicine (2021LK047).

All studies involving human tissues have been approved by the Institutional Ethics Review Committee of Longhua Hospital, affiliated with Shanghai University of Traditional Chinese Medicine (Shanghai, China) (2013LC52), and written informed consent was obtained from the patients in accordance with the Helsinki Declaration. The IACUC number for this animal study is PZSHUTCM221017013. Four-week-old male CAnN.Cg-Foxn1nu/Crl mice were provided with double lion Irradiated Rodent Diet GB 14924.3 and sterile water, and were housed in IVCs mice cage with five mice per cage, under SPF conditions with a 12-h light/dark cycle. The Table of Materials includes detailed information about all materials, reagents, and instruments used in this protocol.
1. Preparation of human osteosarcoma tissue
NOTE: In this study, the human osteosarcoma tissue was resected12 from the femur lesion of a 15-year-old osteosarcoma patient before chemotherapy.
<ol>
	<li>Immediately store the freshly resected osteosarcoma tissue in a tissue-protective solution to maximize the preservation of osteosarcoma cell activation after rinsing with sterile physiological saline. 
	NOTE: Freshly resected osteosarcoma tissues must be transplanted into mice as soon as possible. They can be stored in the tissue protective solution for a maximum of 24 h before transplantation. Osteosarcoma tissues used for modeling must be from patients who have not received chemotherapy. The activity of tumor cells from patients who received chemotherapy will be poor, leading to the failure of model establishment and loss of high fidelity.</li>
	<li>Transfer the freshly resected osteosarcoma tissues stored in the tissue protective solution to the laboratory as soon as possible.</li>
	<li>Prepare experimental instruments and materials for establishing the PDX model: scalpels (Figure 1A); ophthalmic tweezers (Figure 1B,C); ophthalmic scissor (Figure 1D); suture needle (Figure 1E); suture line (Figure 1F); straight needle holder (Figure 1G); marking pen (Figure 1H). 
	NOTE: Ensure that all surgical instruments are sterilized&#160;by autoclaving before use.</li>
	<li>Rinse the osteosarcoma tissues twice with pre-cooled sterile physiological saline in a sterile hood.</li>
	<li>Remove areas of the osteosarcoma tissue with bleeding and necrosis.</li>
	<li>Cut the cleaned osteosarcoma tissue into 3 mm3 pieces with a scalpel in a culture dish containing pre-cooled sterile physiological saline and keep it on ice.</li>
</ol>
2. Establishment of PDX models by osteosarcoma tissue transplantation at mouse flank region
<ol>
	<li>Administer preoperative analgesia of meloxicam (5 mg/kg/24 h) by subcutaneous injection in mice and place the nude mice on sterile surgical drapes. Provide thermal support throughout the procedure.</li>
	<li>Induce anesthesia in mice by exposing them to 3% isoflurane and 97% oxygen. After the mice are completely anesthetized, transfer them to the nose cone and maintain anesthesia with 1.5% isoflurane and 98.5% oxygen.</li>
	<li>Pinch the toes to ensure the mice are completely anesthetized, and wait longer if there are still spasms or convulsive reactions. 
	NOTE: All procedures must be carried out using sterile equipment in a sterile hood. Avoid dryness of mice during anesthesia by applying ophthalmic ointment to their eyes.</li>
	<li>Secure the mouse in a lateral decubitus position (Figure 2A). Disinfect one side of the mouse flank region for osteosarcoma tissue transplantation 3 times with alternating rounds of povidone iodine and 70% ethanol disinfectant cotton balls. 
	NOTE: Surgical disinfection is performed in a circular pattern, beginning in the center and spiraling outward.</li>
	<li>Mark the surgical incision site on the skin with a marker pen.</li>
	<li>Make a 5 mm incision from the skin to the subcutaneous tissues with a scalpel (Figure 2B).</li>
	<li>Lift the upper side skin of the incision margin with ophthalmic tweezers using the left hand, and perform blunt dissection upward under the dermis of the mice with a straight needle holder using the right hand (Figure 2C).</li>
	<li>Hold the upper side skin of the incision with the ophthalmic tweezers using the left hand, and place the osteosarcoma tissue under the skin approximately 5 mm above the incision margin with ophthalmic tweezers using the right hand to transplant the osteosarcoma tissue (Figure 2C). 
	NOTE: Ensure the osteosarcoma tissue is transplanted just below the dermis of the skin. Blunt dissection is performed with a straight and blunt dissection forceps, such as a straight needle holder, to avoid penetrating the mouse thoracic cavity while finding the dermis layer of the mouse skin.</li>
	<li>Suture the incision using absorbable suture with 2-3 stitches for a 5 mm incision (Figure 2D).</li>
	<li>Return the mice to clean cages and monitor them until they completely recover from anesthesia. 
	NOTE: PDX established by transplanting the human osteosarcoma tissue is designated as passage 0 (P0), PDX established by transplanting the PDX tissue at P0 is designated as passage 1 (P1), followed by P2 and P3.</li>
</ol>
3. Collection of PDX tumor tissues
<ol>
	<li>Measure the tumor volume once a week. When the tumor size reaches 1500 mm3, euthanize the mice using the cervical dislocation method after CO2 inhalation. Humane endpoints include tumor ulceration or issues with front limb mobility. 
	NOTE: Measure the long diameter (a) and short diameter (b) of the tumor using a caliper. Calculate the tumor volume (V) using the formula: V = 1/2 &#215; a &#215; b2.</li>
	<li>Position the mouse laterally to expose the tumor, and disinfect the skin at the tumor site with an alcohol-soaked cotton ball.</li>
	<li>Use ophthalmic scissors to separate the entire tumor. Weigh the tumor mass using an electronic scale.</li>
</ol>
4. Pathological examination of primary clinical and PDX tumor tissues
<ol>
	<li>Fix the tumor tissues13 in a 50 mL tube containing 30 mL of 10% neutral buffered formalin solution for 24 h. Rinse the tumor tissues thoroughly with flowing water to remove the fixative.</li>
	<li>Dehydrate and embed the tumor tissues in paraffin14.</li>
	<li>Slice the tumor tissues and conduct the routine histological examination15.</li>
</ol>

Protocol for Osteosarcoma PDX Mouse Models: Studying Growth and Drug Resistance

<ol>
	<li>Saraf, A. J., Fenger, J. M., Roberts, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteosarcoma:+accelerating+progress+makes+for+a+hopeful+future.">Osteosarcoma: accelerating progress makes for a hopeful future.</a> Front Oncol. 8, 4 (2018).</li><li>Ligon, J. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathways+of+immune+exclusion+in+metastatic+osteosarcoma+are+associated+with+inferior+patient+outcomes.">Pathways of immune exclusion in metastatic osteosarcoma are associated with inferior patient outcomes.</a> J Immunother Cancer. 9 (5), e001772 (2021).</li><li>Miwa, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+and+emerging+targets+in+immunotherapy+for+osteosarcoma.">Current and emerging targets in immunotherapy for osteosarcoma.</a> J Oncol. 2019, 7035045 (2019).</li><li>Eaton, B. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteosarcoma.">Osteosarcoma.</a> Pediatr Blood Cancer. 68 (2), e28352 (2021).</li><li>Ritter, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Osteosarcoma.">Osteosarcoma.</a> Ann Oncol. 21, 320-325 (2010).</li><li>Lowery, C. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Broad+spectrum+activity+of+the+checkpoint+kinase+1+inhibitor+prexasertib+as+a+single+agent+or+chemo+potentiator+across+a+range+of+preclinical+pediatric+tumor+models.">Broad spectrum activity of the checkpoint kinase 1 inhibitor prexasertib as a single agent or chemo potentiator across a range of preclinical pediatric tumor models.</a> Clin Cancer Res. 25 (7), 2278-2289 (2019).</li><li>Hingorani, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trastuzumab+deruxtecan,+antibody-drug+conjugate+targeting+Her2,+is+effective+in+pediatric+malignancies:+a+report+by+the+pediatric+preclinical+testing+consortium.">Trastuzumab deruxtecan, antibody-drug conjugate targeting Her2, is effective in pediatric malignancies: a report by the pediatric preclinical testing consortium.</a> Mol Cancer Ther. 21 (8), 1318-1325 (2022).</li><li>Brennecke, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CXCR4+antibody+treatment+suppresses+metastatic+spread+to+the+lung+of+intratibial+human+osteosarcoma+xenografts+in+mice.">CXCR4 antibody treatment suppresses metastatic spread to the lung of intratibial human osteosarcoma xenografts in mice.</a> Clin Exp Metastasis. 31 (3), 339-349 (2014).</li><li>Pan, B., Wei, X., Xu, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patient-derived+xenograft+models+in+hepatopancreatobiliary+cancer.">Patient-derived xenograft models in hepatopancreatobiliary cancer.</a> Cancer Cell Int. 22 (1), 41 (2022).</li><li>Tentler, J. 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S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+practical+protocol+to+prepare+paraffin-embedded+whole+tick+histology+sections.">A practical protocol to prepare paraffin-embedded whole tick histology sections.</a> Ticks Tick Borne Dis. 14 (4), 102162 (2023).</li><li>Chang, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Matrine+inhibits+prostate+cancer+via+activation+of+the+unfolded+protein+response/endoplasmic+reticulum+stress+signaling+and+reversal+of+epithelial+to+mesenchymal+transition.">Matrine inhibits prostate cancer via activation of the unfolded protein response/endoplasmic reticulum stress signaling and reversal of epithelial to mesenchymal transition.</a> Mol Med Rep. 18 (1), 945-957 (2018).</li><li>de Alava, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ewing+sarcoma,+an+update+on+molecular+pathology+with+therapeutic+implications.">Ewing sarcoma, an update on molecular pathology with therapeutic implications.</a> Surg Pathol Clin. 10 (3), 575-585 (2017).</li><li>Hidalgo, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patient-derived+xenograft+models:+An+emerging+platform+for+translational+cancer+research.">Patient-derived xenograft models: An emerging platform for translational cancer research.</a> Cancer Discov. 4 (9), 998-1013 (2014).</li><li>Eoh, K. 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L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+utility+of+patient-derived+xenografts+to+determine+biomarkers+of+prognosis+and+map+resistance+pathways+in+EGFR-mutant+lung+adenocarcinoma.">Clinical utility of patient-derived xenografts to determine biomarkers of prognosis and map resistance pathways in EGFR-mutant lung adenocarcinoma.</a> J Clin Oncol. 33 (22), 2472-2480 (2015).</li><li>Kresse, S. H., Meza-Zepeda, L. A., Machado, I., Llombart-Bosch, A., Myklebost, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preclinical+xenograft+models+of+human+sarcoma+show+nonrandom+loss+of+aberrations.">Preclinical xenograft models of human sarcoma show nonrandom loss of aberrations.</a> Cancer. 118 (2), 558-570 (2012).</li><li>Dobrolecki, L. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patient-derived+xenograft+(PDX)+models+in+basic+and+translational+breast+cancer+research.">Patient-derived xenograft (PDX) models in basic and translational breast cancer research.</a> Cancer Metastasis Rev. 35 (4), 547-573 (2016).</li><li>Abdolahi, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patient-derived+xenograft+(PDX)+models,+applications+and+challenges+in+cancer+research.">Patient-derived xenograft (PDX) models, applications and challenges in cancer research.</a> J Transl Med. 20 (1), 206 (2022).</li><li>Deng, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deep+whole-genome+sequencing+identifies+recurrent+genomic+alterations+in+commonly+used+breast+cancer+cell+lines+and+patient-derived+xenograft+models.">Deep whole-genome sequencing identifies recurrent genomic alterations in commonly used breast cancer cell lines and patient-derived xenograft models.</a> Breast Cancer Res. 24 (1), 63 (2022).</li><li>Chen, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+and+characterization+of+patient-derived+xenografts+from+patients+with+osteosarcoma.">Generation and characterization of patient-derived xenografts from patients with osteosarcoma.</a> Tissue Cell. 79, 101911 (2022).</li><li>Fortuna-Costa, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+association+between+successful+engraftment+of+osteosarcoma+patient-derived+xenografts+and+clinicopathological+findings.">An association between successful engraftment of osteosarcoma patient-derived xenografts and clinicopathological findings.</a> Histol Histopathol. 35 (11), 1295-1307 (2020).</li><li>Beck, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Canine+and+murine+models+of+osteosarcoma.">Canine and murine models of osteosarcoma.</a> Vet Pathol. 59 (3), 399-414 (2022).</li></ol>

The authors declare no competing financial interests.

The PDX models can simulate the characteristics of human cancers and retain more similarity with the primary tumor, including genetic and genomic alterations, histology, heterogeneity, and gene expression profile16,17,18,19. Therefore, they preserve the molecular phenotypes and genotypes of cancer patients, providing innovative approaches for studying biology and evaluating potential therapeuti...

Osteosarcoma is a primary bone malignancy derived from interosseous lobe cells and is most common in adolescents as well as children. It often occurs in the epiphysis of the long diaphysis and is characterized by high malignancy, early metastasis, and poor prognosis1,2. Lung metastasis is the main cause of death in osteosarcoma patients. The 5-year survival rate of patients with non-metastatic osteosarcoma is 65%-70%3. However, over the last 40 years, the 5-year survival rate (only 20%) of patients with metastatic osteosarcoma has not significantly improved, and 25% of osteosarcoma patients have metastases at the time of diagnosis4. Currently, the first-line drugs for osteosarcoma treatment have reached a consensus, but there are still significant differences in chemotherapy regimen and treatment time5. It is important to perform preclinical experiments based on appropriate animal models to obtain more effective chemotherapy regimens.
Currently, models commonly used for osteosarcoma preclinical experiments include cell line-based in vitro cell culture and in vivo cell-derived xenografts (CDX), as well as patient-derived xenografts (PDX)6,7.
The cell lines are convenient for culturing and for use in in vitro studies, or for transplantation into immunodeficient mice to establish CDX models8. However, cell lines cultured in vitro may not accurately reflect the heterogeneity of malignancies and the individual characteristics of patients due to potential mutations that occur to adapt to the in vitro culture environment during repeated passages. Additionally, they lack the microenvironment and immune system necessary for tumor growth and development in vivo. While CDX models offer some advantages over in vitro cell culture, they still may not fully reflect the individual characteristics of osteosarcoma patients, although tumor tissues obtained from CDX models have limited intratumoral heterogeneity and immune system representation compared to cell lines cultured in vitro9. Therefore, establishing a preclinical model with high fidelity is crucial.
PDX models involve the immediate transplantation of freshly resected human cancer tissues into immunodeficient mice. This method allows for the faithful preservation of genetic, epigenetic, and heterogeneous characteristics of human malignancies for each patient, even after successive passages in mice. Furthermore, PDX models are known to accurately predict later clinical outcomes10, making them valuable tools for creating individualized treatments and advancing precision medicine research11.
This work describes the procedure for establishing a PDX model in immunodeficient mice by transplanting human osteosarcoma tissue. Such models serve as platforms for conducting preclinical experiments for osteosarcoma.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>10% formalin neutral solution</td>
			<td>Wuhan Saiweier Biotechnology Co., Ltd</td>
			<td>G1101-500ml</td>
			<td>Fix the tissues</td>
		</tr>
		<tr>
			<td>Autoclave</td>
			<td>Japan Hiryama Company</td>
			<td>HVE-50</td>
			<td>Sterilization surgical instruments</td>
		</tr>
		<tr>
			<td>CAnN.Cg-Foxn1nu/Crl</td>
			<td>Shanghai SLAC Laboratory Animal Co, Ltd.</td>
			<td>/</td>
			<td>Animal</td>
		</tr>
		<tr>
			<td>Caliper</td>
			<td>Yantai Green Forest Tools Co., Ltd.</td>
			<td>034180A</td>
			<td>Measure the tumor volume</td>
		</tr>
		<tr>
			<td>Dish (60mm)</td>
			<td>Shanghai NianYue Biotechnology Co., Ltd</td>
			<td>430166, Corning</td>
			<td>Sample placment during transplantation</td>
		</tr>
		<tr>
			<td>Disinfectant cotton balls</td>
			<td>Shanghai Honglong Industrial Co., Ltd.</td>
			<td>20230627</td>
			<td>Disinfect the skin of mice</td>
		</tr>
		<tr>
			<td>Disposable sterile gloves</td>
			<td>Guilin Hengbao Health Protection Co., Ltd.</td>
			<td>YT21131</td>
			<td>Sterile operation</td>
		</tr>
		<tr>
			<td>Double lion Irradiated Rodent Diet</td>
			<td>Suzhou Shuangshi Experimental Animal Feed Technology Co., Ltd.</td>
			<td>GB 14924.3</td>
			<td>Animal feed</td>
		</tr>
		<tr>
			<td>Electronic scale</td>
			<td>Shanghai NianYue Biotechnology Co., Ltd</td>
			<td>1-2000</td>
			<td>Weigh the weight of the tumor</td>
		</tr>
		<tr>
			<td>Eosin</td>
			<td>Shanghai Gengyun Biotechnology Co., Ltd</td>
			<td>E4009-25G</td>
			<td>Hematoxylin eosin stain</td>
		</tr>
		<tr>
			<td>Hematoxylin</td>
			<td>Shanghai Gengyun Biotechnology Co., Ltd</td>
			<td>H3136-25G</td>
			<td>Hematoxylin eosin stain</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Shenzhen RWD Life Technology Co., Ltd</td>
			<td>VETEASY</td>
			<td>Mouse anesthesia&#160;</td>
		</tr>
		<tr>
			<td>IVCs mice cage</td>
			<td>Suzhou Monkey King Animal Experimental Equipment Technology Co., Ltd.</td>
			<td>HH-MMB-2</td>
			<td>Animal barrier</td>
		</tr>
		<tr>
			<td>Mark pen</td>
			<td>Zebra Trading (Shenzhen) Co., Ltd.</td>
			<td>YYST5</td>
			<td>Mark the surgical incision</td>
		</tr>
		<tr>
			<td>Olympus Optical microscope</td>
			<td>Japanese Olympus Company</td>
			<td>BH20</td>
			<td>Scan tissue slices</td>
		</tr>
		<tr>
			<td>Ophthalmic ointment</td>
			<td>Shanghai Gengyun Biotechnology Co., Ltd</td>
			<td>SOICOEYEGRL</td>
			<td>Avoid dry eyes of mice during anesthesia</td>
		</tr>
		<tr>
			<td>Ophthalmic scissors</td>
			<td>Shanghai NianYue Biotechnology Co., Ltd</td>
			<td>Y00030 JZ</td>
			<td>Cut the skin</td>
		</tr>
		<tr>
			<td>Ophthalmic tweezers</td>
			<td>Shanghai NianYue Biotechnology Co., Ltd</td>
			<td>BS-ZER-S-100 Biosharp</td>
			<td>Hold osteosarcoma tissues during transplantation</td>
		</tr>
		<tr>
			<td>Paraffin</td>
			<td>Jiangsu Shitai Experimental Equipment Co., Ltd.</td>
			<td>80200-0015</td>
			<td>Buried osteosarcoma tissue</td>
		</tr>
		<tr>
			<td>Paraffin slicing machine</td>
			<td>Lyca Microsystem (Shanghai) Trading Co., Ltd.</td>
			<td>RM2235</td>
			<td>Osteosarcoma tissue section</td>
		</tr>
		<tr>
			<td>physiological saline</td>
			<td>Guangzhou Jinsheng Biotechnology Co., Ltd.</td>
			<td>605-004057</td>
			<td>Rinse and temporary storage of osteosarcoma tissue</td>
		</tr>
		<tr>
			<td>Scalpels</td>
			<td>Surgical Instrument Factory of Shanghai Medical Devices (Group) Co., Ltd.</td>
			<td>J11010-10# JZ</td>
			<td>Separation of osteosarcoma tissue and making surgical incisions</td>
		</tr>
		<tr>
			<td>Sterile hood</td>
			<td>Thermo Fisher Technology (China) Co., Ltd.</td>
			<td>ECO0.9</td>
			<td>Surgical operation table</td>
		</tr>
		<tr>
			<td>sterile surgical drapes</td>
			<td>Henan Huayu Medical Equipment Co., Ltd.</td>
			<td>20160090</td>
			<td>Provide sterile surgery area</td>
		</tr>
		<tr>
			<td>Straight needle holder</td>
			<td>Shanghai Gengyun Biotechnology Co., Ltd</td>
			<td>J31050 JZ</td>
			<td>Suture the wound</td>
		</tr>
		<tr>
			<td>Suture line</td>
			<td>Shanghai Pudong Jinhuan Medical Products Co., Ltd</td>
			<td>F3124</td>
			<td>Suture the wound</td>
		</tr>
		<tr>
			<td>Suture needle</td>
			<td>Shanghai Pudong Jinhuan Medical Products Co., Ltd</td>
			<td>F3124</td>
			<td>Suture the wound</td>
		</tr>
		<tr>
			<td>Tissue protective solution</td>
			<td>Nanjing Shenghang Biotechnology Co., LTD</td>
			<td>BC-CFM-03</td>
			<td>Maintain the activity of tissue cells</td>
		</tr>
		<tr>
			<td>Tube (50 mL)</td>
			<td>Shanghai Baisai, Biotechnology Co., Ltd.</td>
			<td>BLD-BL2002500</td>
			<td>Install formalin fixation solution</td>
		</tr>
	</tbody>
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establishment of patient-derived xenograft mouse model with human osteosarcoma tissues

Osteosarcoma is the most common primary malignant bone tumor in children and adolescents. Despite the development of new treatment plans in recent years, the prognosis for osteosarcoma patients has not significantly improved. Therefore, it is crucial to establish a robust preclinical model with high fidelity. The patient-derived xenograft (PDX) model faithfully preserves the genetic, epigenetic, and heterogeneous characteristics of human malignancies for each patient. Consequently, PDX models are considered authentic in vivo models for studying various cancers in transformation studies. This article presents a comprehensive protocol for creating and maintaining a PDX mouse model that accurately mirrors the morphological features of human osteosarcoma. This involves the immediate transplantation of freshly resected human osteosarcoma tissue into immunocompromised mice, followed by successive passaging. The described model serves as a platform for studying the growth, drug resistance, relapse, and metastasis of osteosarcoma. Additionally, it aids in screening the target therapeutics and establishing personalized treatment schemes.

Author Spotlight: Replicating Human Osteosarcoma Progression in Immunodeficient Mice for Cancer Study

Establishment of Patient-Derived Xenograft Mouse Model with Human Osteosarcoma Tissues

This study meticulously replicates the lateral progression of human osteosarcoma tissue growth in immunodeficient mice, providing a platform for studying the growth, the drug resistance, relapse, and metastasis of osteosarcoma. We realized the procedural steps involved in establishing a patient-derived xenograft model using human osteosarcoma tissue, thereby advising the preclinical research on osteosarcoma. This technical accuracy mimics the human cancer and maintains primary tumor similarity, facilitating the exploration of osteosarcoma biology and the treatment efficacy.

This protocol describes the detailed procedure for establishing a PDX mouse model, preserving the morphological features of human osteosarcoma after immediate transplantation of freshly resected human osteosarcoma tissue and successive passages in mice. Here, a PDX mouse model was successfully established using human osteosarcoma tissue.
Figure 3A shows a representative mouse of PDX at P0, two months after the transplantation of femoral osteosarcoma tissue from a ...

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The present protocol describes the method for establishing a patient-derived xenograft (PDX) mouse model using human osteosarcoma tissue.

Osteosarcoma is the most common primary malignant bone tumor in children and adolescents. Despite the development of new treatment plans in recent years, ...

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Research

JoVE Journal

Cancer Research

418 Views.  Shanghai University of Traditional Chinese Medicine. This study meticulously replicates the lateral progression of human osteosarcoma tissue growth in immunodeficient mice, providing a platform for studying the growth, the drug resistance, relapse, and metastasis of osteosarcoma. We realized the procedural steps involved in establishing a patient-derived xenograft model using human osteosarcoma tissue, thereby advising the preclinical research on osteosarcoma. This technical accuracy mimics the human cancer and maintains primary tumor similarit...

Video: Author Spotlight: Replicating Human Osteosarcoma Progression in Immunodeficient Mice for Cancer Study

Establishment of PDX Models by Osteosarcoma Tissue Transplantation at the Mouse Flank Region

To begin, take the freshly resected osteosarcoma tissue, stored in the tissue protective solution. Rinse the osteosarcoma tissues twice with pre-cool sterile physiological saline in a sterile hood. Then remove the areas of osteosarcoma tissue with bleeding and necrosis.Using a scalpel, cut the clean osteosarcoma tissue in a tissue culture dish containing pre-cool sterile physiological saline. Keep the tissue on ice. Now, place the nude mice on sterile surgical drapes.Pinch the toes of the mice to ensure that they are completely anesthetized. Then secure the mouse in a lateral decubitus position. With a marker pen, mark the surgical incision site on the skin.Use the scalpel to make a five millimeter incision from the skin to the subcutaneous tissues. Now, hold ophthalmic tweezers in the left hand and lift the upper margin of the incision. Simultaneously, use a straight needle holder with the right hand to perform blunt dissection upward under the dermis of the mice.With ophthalmic tweezers, hold the upper side skin of the incision. Then transplant the osteosarcoma tissue under the skin five millimeters above the incision margin. Suture the incision using surgical lines, with two to three stitches for a five millimeter incision.Transplanting femoral osteosarcoma tissue from a 15-year-old patient to P0, P1, P2, and P3 mice resulted in the successful creation of a patient-derived xenograft.