RB1 is inactivated in UPN-1 and in the original primary MCL. (A) Multiplex PCR amplifications of RB1 exons 1, 17, and 18 performed with genomic DNA from UPN-1 cells are shown. Although β-actin (β-act) was amplified in both UPN-1 and in the control (C+), only exon 18 of RB1 was amplified in UPN-1. (B) A section of the Log2Ratio graph corresponding to the 13q14 region generated by the CNAT application is showed. All SNPs are displayed in blue color indicating that the log2 ratio values were higher than 0. A zoom view of a small region containing negative log2 ratio values show that it correspond to the RB1 locus. Two maps are shown, one represents all RB1 exons by gray vertical lines, and the second represents the SNPs located in the region. In red are indicated the SNPs that are deleted with signal log2 ratio value lower than −1, and conserved SNPs are shown in black. (C) pRB1 expression was analyzed by Western blot in 5 cell lines. Equal amount of total protein was loaded in each lane. A section of the Coomassie staining gel is showed at the bottom. (D) Immunohistochemical staining of pRB1 in the primary tumor from which UPN-1 was established. Tumor cells are negative, whereas endothelial cells, considered as internal positive controls, showed positivity (inset). (E) A multiplex PCR reaction was designed to amplify exon 1 and 18 of RB1 together with β-actin. Both RB1 exons were amplified when we used a DNA control (C+), but when we used DNA extracted from the primary tumor from which the UPN-1 cell line was established (UPN-1 case) only the RB1 exon 18 was amplified.