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Ble cross-hybridizing host genes. The use of our animal model to identify mediators of glioma invasion has the potential pitfall of identifying artifacts of xenografting. That is, human glioma cells confronted with nude mouse brain rather than human brain may express genes specific to this setting. Two arguments can be made against this theory. First, there is no teleological reason for human cell
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Radial migration assay was performed as previously described [30,31]. In a blinded fashion, various U87Gal-1 clones were analyzed and compared to U87GFP controls and parental U87MG cells. Of each clone, 2500 cells were allowed to sediment through a cooled manifold onto laminin-coated cell culture wells (Creative Scientific Methods, Inc., Phoenix, AZ). The manifold and slide were incubated together
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Orithm. Galectin-1 was ranked at the top of this list. We thus took advantage of our own patient-derived glioblastoma xenograft model [25] in order to further decipher the roles of galectin-1 on GBM cell migration features. The system we have developed mitigates the effect of three important confounders from human samples. First, tissue is frozen within one minute of removal, ensuring high quality
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Ss microarray chip platforms when hybridizing the same sample RNA [32]. Additionally,Toussaint et al. Molecular Cancer 2012, 11:32 http://www.molecular-cancer.com/content/11/1/Page 8 ofFigure 4 (A) Galectin-1 transfection does not alter U87MG attachment to fibronectin. Attachment to fibronectin-coated 96-well plates was quantitated with an MTT assay with and without a media change in the middle of
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Ns were transferred to nitrocellulose and these membranes were incubated with primary antibody for 60 minutes (anti-Gal1 from Research Diagnostics, Flanders, New Jersey, anti-beta actin from Sigma, St. Louis, Missouri). After washing and incubation with secondary antibody (Goat Anti-Mse IgG-HRP, Pierce, Rockford, IL), developing solution was added to the membrane (Supersignal West Femto Substrate,
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Few of our U87 galectin1 clones. Parental U87MG cells, along with galectin-1 and acGFP-only clones were injected into the right caudate/putamen complex of nude mice. Tumors overexpressing galectin-1 shortened survival of their hosts compared to their parental counterparts (Figure 5). A few animals (7/20) bearing tumors expressing acGFP alone eventually exhibited neurological symptoms. The examinat