Where Ideas Grow

In vivo CAM assays

The In vivo CAM assays platform provides scientific expertise and services using of the chick embryo model, more specifically, assays based on the chorioallantoic membrane – the CAM.

Created in 2012, the platform offers researchers additional or alternative in vivo tools (complying with the 3Rs policy) that are reliable and cost and time efficient.

Due to the CAM structure and its easy access, CAM assays constitute attractive preclinical in vivo tools for drug screening and/or vascular growth studies. Associated with the chick natural immunoincompetence, CAM assays can also be used to study complex cancer features and the effect of potential therapeutic molecules.

The platform ensures:

  • Protocol design
  • Experimental execution
  • Analysis of the results
  • Data interpretation.

 Assays are available for all scientific community to analyze functional features such as:

  • Angiogenesis
  • Tumorigenesis
  • Cell Invasion
  • Metastisation
  • Vascular Permeability

The available collection of functional assays can be applied to a diversity of test conditions:

  • Cells previously grown in culture
  • Biomaterials assembled with or without cells
  • Drugs, tested directly on the CAM or on CAM xenograft tumours
  • Conditioned mediums
  • Supernatants enriched in exosomes or other vesicles or components
  • Extracts from plants or microorganisms.

The platform is exclusively dedicated to CAM assays and is fully equipped to address new challenges and widen the model applications in a range of research topics. The In vivo CAM assays platform’s work is validated by our publication track record, and by the national and international network of collaborators and clients, both from academia and industry.

Platform Head



In vivo analysis of functional features:

  • Angiogenesis (e.g.)
  • Tumourigenesis (e.g.)
  • Cell Invasion (e.g.)
  • Metastisation 
  • Vascular Permeability (e.g.)



Multiple test conditions:

  • cells previously grown in culture (normal or tumour)
  • biomaterials (assembled with or without cells)
  • drugs, tested directly on the CAM (anti- or pro- angiogenic activity)
  • drugs, tested on CAM xenograft tumours (anti- or pro- tumourigenic activity)
  • conditioned mediums (from cultured cells)
  • supernatants enriched in components of interest (exosomes, etc)
  • extracts from plants or microorganisms or derived proteins
  • etc



The chick embryo has long been used as a model organism in a number of areas including oncobiology and biomaterial research. The embryo is surrounded by the chorioallantoic membrane (CAM), a highly vascularised extra-embryonic membrane that can be used to graft human cells. When grafted on CAM, tumour cells are capable of stimulating the formation of new blood vessels, benefiting from blood supply. This allows them to develop in a similar manner as in their native microenvironment i.e. to proliferate, invade and metastasise to the chick embryonic organs.

The in vivo chick embryo model presents several important advantages:

  • It fulfils the 3Rs policy.  Both the Directive 2010/63/EU of the European Parliament, and the Portuguese law DL 113/2013,  on the protection of animals used for scientific purposes, do not contain any kind of restriction to the use of non-mammal embryos
  • The embryo is naturally immunoincompetent, thus easily allowing mammal cell xenografts
  • It requires short experimental times


Publications with the participation of the i3S in vivo CAM Assays Platform:

  • MT Pinto, AS Ribeiro, I Conde, R Carvalho, J Paredes (2021) The Chick Chorioallantoic Membrane Model: A New In Vivo Tool to Evaluate Breast Cancer Stem Cell Activity. Int. J. Mol. Sci. 2021,22, 334.https://doi.org/10.3390/ijms22010334 (JIF:4.556)


  • M Dionisio, A Vieira, R Carvalho, I Conde, M Oliveira, M Gomes, M Pinto, P Pereira, J Pimentel, C Souza, M Silveira, V da Silva, A Barroso, D Preto, J Cameselle-Teijeiro, F Schmitt, A Ribeiro and J Paredes (2020)  BR-BCSC Signature: The Cancer Stem Cell Profile Enriched in Brain Metastases that Predicts a Worse Prognosis in Lymph Node-Positive Breast Cancer. Cells 2020, 9(11), 2442https://doi.org/10.3390/cells9112442 (JIF:4.829)


  • C Coelho, T Padrão, L Costa, M Pinto, P Costa, V Domingues, P Quadros, F Monteiro, S Sousa. (2020)The antibacterial and angiogenic effect of magnesium oxide in a hydroxyapatite bone substitute. Sci Rep 10, 19098 (2020). https://doi.org/10.1038/s41598-020-76063-9 (JIF:4.120)


  • M Leite, MS Marques, J Melo, MT Pinto, B Cavadas, M Aroso, M Gomez-Lazaro, R Seruca, C Figueiredo. Helicobacter Pylori Targets the EPHA2 Receptor Tyrosine Kinase in Gastric Cells Modulating Key Cellular Functions" Cells. 2020 Feb 24;9(2). pii: E513. doi: 10.3390/cells9020513. (JIF:4.829)

  • A S. Silva, L F. Santos, MC. Mendes and JF. Mano (2020) Multi-layer pre-vascularized magnetic cell sheets for bone regeneration. Biomaterials, Volume 231, 119664, ISSN 0142-9612. https://doi.org/10.1016/j.biomaterials.2019.119664.

  • AR Sousa, C Martins?Cruz, MB. Oliveira and JF. Mano (2020) One?Step Rapid Fabrication of Cell?Only Living Fibers Adv. Mater. 2020, 32, 1906305. https://doi.org/10.1002/adma.201906305

  • Alves MM, Mil-Homens D, Pinto S, Santos CF, Montemor MF. Antagonist biocompatibilities of Zn-based materials functionalized with physiological active metal oxides. Colloids Surf B Biointerfaces. 2020;191:110990. doi:10.1016/j.colsurfb.2020.110990 (JIF: 4.389)

  • Coelho R, Ricardo S, Amaral AL, et al. Regulation of invasion and peritoneal dissemination of ovarian cancer by mesothelin manipulation. Oncogenesis. 2020;9(6):61. Published 2020 Jul 1. doi:10.1038/s41389-020-00246-2 (JIF: 5.995)

  • Ghazaryan N, Movsisyan N, Macedo JC, et al. The antitumor efficacy of monomeric disintegrin obtustatin in S-180 sarcoma mouse model. Invest New Drugs. 2019;37(5):1044-1051. doi:10.1007/s10637-019-00734-2 (JIF: 3.525)

  • CR. Correia, IM. Bjørge, J Zeng, M Matsusaki and JF. Mano (2019) Liquefied Microcapsules as Dual?Microcarriers for 3D+3D Bottom?Up Tissue Engineering. Adv. Healthcare Mater. 8, 1901221. https://doi.org/10.1002/adhm.201901221 

  • Janser, FA, Ney, P, Pinto, MT, Langer, R and Tschan, MP (2019). The Chick Chorioallantoic Membrane (CAM) Assay as a Three-dimensional Model to Study Autophagy in Cancer Cells. Bio-protocol 9(13): e3290. DOI: 10.21769/BioProtoc.3290.

  • JC Loureiro, AL Torres, T Neto, P Aguiar, C Barrias, MT Pinto, I.F. Amaral. Conjugation of the T1 sequence from CCN1 to fibrin hydrogels for therapeutic vascularization (2019) Materials Science & Engineering C Volume 104, November 2019, 109847. https://doi.org/10.1016/j.msec.2019.109847 (JIF: 5.08)

  • FG Marques, E Poli, J Rino, MT Pinto, I Diegues, F Pina, S Constantino Rosa Santos. Low doses of ionizing radiation enhance the angiogenic potential of adipocyte conditioned medium. Radiat Res. 2019 Nov;192(5):517-526. doi: 10.1667/RR15438.1. (JIF:2.539)

  • D Freitas, D Campos, J Gomes, F Pinto, JA Macedo, R Matos, S Mereiter, MT Pinto, A Polónia, F Gartner, C Steentoft, A Magalhães, CA Reis. (2019) O-glycans truncation modulates gastric cancer cell signalling and transcription leading to a more aggressive phenotype. EBioMedicine. 2019 Feb;40:349-362. doi: 10.1016/j.ebiom.2019.01.017 (JIF:6.18)

  • M Santos, PM Pereira, AS Varanda, J Carvalho, M Azevedo, DD Mateus, N Mendes, P Oliveira, F Trindade, MT Pinto, R Bordeira-Carriço, F Carneiro, R Vitorino, C Oliveira, MA Santos. (2018) Codon misreading tRNAs promote tumor growth in mice. RNA Biol. Mar 20:1-38. doi: 10.1080/15476286.2018.1454244 (JIF:3.9)

  • AL Torres, SJ Bidarra, MT Pinto, C Aguiar, EA Silva, C Barrias. (2018) Guiding morphogenesis in cell-instructive microgels for therapeutic angiogenesis. Biomaterials. Vol 154, Feb 2018,  34-47. doi: 10.1016/j.biomaterials.2017.10.051  (JIF:8.402)

  • D Olivera-Severo, AF Uberti, MS Marques, MT Pinto, M  Gomez-Lazaro, C Figueiredo, M  Leite, CR Carlini. (2017) A New Role for Helicobacter pylori Urease: Contributions to Angiogenesis. Frontiers in Microbiology. Vol8, Nº 1883. doi:  10.3389/fmicb.2017.01883 (JIF:4.076)

  • H Pópulo, B Nunes, C Sampaio, R Baptista, MT Pinto, TB Gaspar, LM Alves, R Cai, XY Zhang,  AV Schally M Sobrinho-Simões, P  Soares. (2017) Inhibitory effects of antagonists of growth hormone releasing hormone (GHRH) in thyroid cancer. Hormones and Cancer, Sep 18. doi: 10.1007/s12672-017-0307-4 (JIF:2.93)

  • TM Ribeiro-Rodrigues, TL Laundos, R. Pereira-Carvalho, D Batista-Almeida, R Pereira, V Coelho-Santos, AP Silva, R Fernandes, M Zuzarte, F  Enguita, MC Costa, P Pinto-do-Ó, MT Pinto, P Gouveia, L Ferreira, JC Mason, P Pereira, B Kwak, D Nascimento and H Girão. (2017)  Exosomes secreted by cardiomyocytes subjected to ischemia promote cardiac angiogenesis. Cardiovascular Research. 113(11):1338-1350. doi: 10.1093/cvr/cvx118 (JIF:5.88)

  • AT Pinto, ML Pinto, S. Velho,  MT Pinto, AP Cardoso, R. Figueira, A Monteiro, M Marques, R Seruca, MA Barbosa, M Mareel and MJ Oliveira.(2016) Intricate macrophage-colorectal cancer cell communication in response to radiation. PlosOne, 2016 Aug 11; 11(8):e0160891. doi: 10.1371/journal.pone.0160891 (JIF: 3.54)

  • LB Ferreira, C Tavares, A Pestana, CL Pereira, C Eloy, MT Pinto, P Castro, R Batista, E Rios, M Sobrinho-Simões, ER Gimba, P Soares.(2016) Osteopontin-a splice variant is overexpressed in papillary thyroid carcinoma and modulates invasive behavior. Oncotarget. 2016, 7(32):52003-52016. doi: 10.18632/oncotarget.10468 (JIF:6.36)

  • AT Pinto, ML Pinto, AP Cardoso, C Monteiro, MT Pinto, A Maia, P Castro, R Figueira, A Monteiro, M Marques, M Mareel, SG Santos, R Seruca, MA Barbosa, S Rocha and MJ Oliveira. (2016) Ionizing radiation modulates human macrophages towards a pro-inflammatory phenotype preserving their pro-invasive and pro-angiogenic capacities. Scientific Reports, Jan 6;6:18765.  doi: 10.1038/srep18765. (JIF: 5.58)

  • MF Estrada, S Rebelo, E Davies, MT Pinto, H Pereira, VE Santo, MJ Smalley, ST Barry, E Gualda, PM Alves, E Anderson, C Brito.(2016) Modelling the tumour microenvironment in long-term microencapsulated 3D co-cultures recapitulates phenotypic features of disease progression. Biomaterials, Feb; 78:50-61. doi: 10.1016/j.biomaterials.2015.11.030. (JIF: 8.56)

  • MI Almeida, A Machado Silva, D Marques Vasconcelos, CR Almeida, H Caires, MT Pinto, GA Calin, S Gomes Santos, MA Barbosa. (2016) miR-195 in human primary Mesenchymal Stromal/Stem Cells regulates proliferation, osteogenesis and paracrine effect on angiogenesis. Oncotarget, 7(1):7-22 doi: 10.18632/oncotarget.6589. (JIF: 6.63)

  • C Resende, G Regalo, C Durães, MT Pinto, W Xiaogang, C Figueiredo, F Carneiro, JC Machado. (2016) Interleucin-1B signaling leads to increased cell survival of gastric carcinoma cells through a CREB/CEBPß-associated mechanism. Gastric Cancer, Jan;19(1):74- 84. doi: 10.1007/s10120-014-0448-x (JIF:4.828)

  • J Caldeira, J Figueiredo, C Brás-Pereira, MT Pinto, JB Relvas, M Barbosa, F Casares, F Janody, R Seruca. (2015) E-cadherin-defective Gastric Cancer Cells Depend on Laminin to Survive and Invade. Human Molecular Genetics 24(20):5891-900. doi:10.1093/hmg/ddv312. (JIF:6.39)

  • AP Cardoso, ML Pinto, AT Pinto, MT Pinto, C Monteiro, MI Oliveira, SG Santos, JB Relvas, R Seruca, A Mantovani, M Mareel, MA Barbosa, MJ Oliveira. (2015) Matrix metalloproteases as maestros for the dual role of LPS- and IL-10-stimulated macrophages in cancer cell behavior. BMC cancer 15:456. doi: 10.1186/s12885-015-1466-8 (JIF: 3.32)

  • AP Cardoso, ML Pinto, AT Pinto, MI Oliveira, MT Pinto, R Gonçalves, JB Relvas, C Figueiredo, R Seruca, A Mantovani, M Mareel, MA Barbosa, MJ Oliveira (2014) Macrophages stimulate gastric and colorectal cancer invasion through EGFR Y(1086), c-Src, Erk1/2 and Akt phosphorylation and smallGTPase activity. Oncogene 33(16):2123-33. doi: 10.1038/onc.2013.154. (JIF:7.36)

  • J Simões-Correia , DI Silva, S Melo, J Figueiredo, J Caldeira, MT Pinto, H Girão, P Pereira, R Seruca (2014) DNAJB4 molecular chaperone distinguishes WT from mutant E-cadherin, determining their fate in vitro and in vivo. Human Molecular Genetics Apr 15; 23(8):2094- 105. doi: 10.1093/hmg/ddt602 (JIF:7.69)

  • C Gomes, H Osório, MT Pinto, D Campos, MJ Oliveira, C Reis (2013) Expression of ST3GAL4 Leads to SLe(x) Expression and Induces c-Met Activation and an Invasive Phenotype in Gastric Carcinoma Cells. PLOsOne vol 8: 6, e66737. doi:10.1371/journal.pone.0066737. (JIF:3.7)

  • H Pinheiro, J Carvalho, P Oliveira, D Ferreira, MT Pinto, H Osório, D Licastro, R Bordeira- Carriço, P Jordan, D Lazarevic, R Sanges, E Stupka, D Huntsman, R Seruca, C Oliveira. (2012) Transcription initiation arising from E-cadherin/CDH1 intron2: a novel protein isoform that increases gastric cancer cell invasion and angiogenesis. Human Molecular Genetics Vol. 21,No. 19, 4253–4269 . doi: 10.1093/hmg/dds248 (JIF:7.69)

  • J Caldeira, J Simões-Correia, J Paredes, MT Pinto, S Sousa, G Corso, R Roviello, P Pereira, D Weil, C Oliveira, F Casares, R Seruca. (2012) CPEB1, a novel gene silenced in gastric cancer: a Drosophila approach. Gut Aug;61(8):1115-23. doi: 10.1136/gutjnl-2011-300427 (JIF:10,6)

  • M Pinto, P Soares, D Ribatti.(2011) Thyroid hormone as a regulator of tumor induced angiogenesis. Cancer Letters Feb 28; 301(2):119-26. doi: 10.1016/j.canlet.2010.11.011 (JIF:4.2)

  • H Prazeres, J Torres, F Rodrigues, M Pinto, MC Pastoriza , D Gomes, J Cameselle- Teijeiro, A Vidal, T Martins, M Sobrinho-Simões, P Soares (2011) Chromosomal, epigenetic and microRNA-mediated inactivation of LRP1B, a modulator of the extracellular environment of thyroid cancer cells. Oncogene Mar 17; 30(11):1302-17. doi: 10.1038/onc.2010.512 (JIF:7.2)


DIRECTIVE 2010/63/EU OF THE EUROPEAN PARLIAMENT AND OF THE COUNCIL of 22 September 2010 on the protection of animals used for scientific purposes


Decreto-Lei n.º 113/2013


i3S Scientific Platforms Acknowledgements