Y one-dimensional gel electrophoresis for direct comparison of the protein level in the different cell

Y one-dimensional gel electrophoresis for direct comparison of the protein level in the different cell types on the same immunoblot (d). All immunoblots shown are representative for three or more performed experiments. See Material and Methods for details on cell separation and immunoblotting technique.Page 6 of(page number not for citation purposes)BMC Bioinformatics 2006, 7:http://www.biomedcentral.com/1471-2105/7/Correlation of p53 protein isoforms towards the AML differentiation level The French-American-British (FAB) Olumacostat glasaretil biological activity classification of AML is based on the morphologically determined stage of myeloid maturation and direction of maturation [36,37]. Recent reports indicate that the FAB classification, in particular the distinction between M1? and M4? in maturation level and direction of maturation, is associated with certain gene classes in unsupervised clustering of gene expression profiles [38,39]. It is previously described in several reports that p53 is involved in leukemic cell differentiation [24,25,40,41]. Phosphorylation of p53 Ser315 is necessary for differentiation in mouse embryonic stem cells [42], and p53 is able to direct differentiation in AML cell lines [25,41]. The p53-deficient HL-60 cell line has potential for both monocytic and granulocytic differenti-ation, and introduction of wild type p53 directs differentiation in the granulocytic direction [40]. Based on these reports we hypothesized that the p53 biosignatures should reflect the stage and direction of myeloid differentiation. Therefore, we measured correlations between the established routine morphological differentiation classification of AML (FAB) [21,22,35] and the p53 2DE biosignatures of the cancer cells. We assigned to every class a separate t-value: M0 (t = 0), M1 (t = 1), M2 (t = 2), M3 (t = 3), M4 (t = 4) and M5 (t = 5). Using 73 gels we found specific correlations (Fig. 4). Image A is the masked correlation landscape, image B is the raw correlation image. The observations were: a) The tail of the p53- isoform correlates negatively to the FAB classification (profile 4, region g and h). b) The p63 areaFigure 4 Correlation of p53 protein isoforms towards AML differentiation (FAB) Correlation of p53 protein isoforms towards AML differentiation (FAB). Green indicates correlation with more differentiated forms of AML. Such areas in 2DE images of M5 will have a higher intensity than 2DE images of M0. Brown indicates anti-correlation with the more mature forms of leukemia cells. Such areas in 2DE images of M0 will have a higher intensity than 2DE images of M5. (A) Correlation landscape of p53 in 73 AML images related to differentiation direction and stage (FAB, French-American-British classification). The vertical axis sets out the absolute correlation value. (B) Correlation image demonstrating statistical significant alterations in p53. Profile 1 shows the p53- region containing four correlating spots (r = 0.2). Profile 2 shows the sub- region anti-correlating at positions e and f. Profile 3 is the p63 region (p53 family member) correlating towards the more differentiated leukemia’s. Profile 4, a p53 region anti-correlating with differentiated AML.Page 7 of(page number not for citation purposes)BMC Bioinformatics 2006, 7:http://www.biomedcentral.com/1471-2105/7/correlates positively towards PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/27735993 the FAB classification (profile 3, the i region), c) The p53- region has four positively correlating articulated spots (profile 1, a-d, r = 0.2), d) the p53 sub- region ha.