Conventional RT-PCR and direct cDNA sequencing confirm the presence of <i>EGFRvIII</i> in glioblastoma FFPE tissue.

<p>A. Conventional RT-PCR failed for GBM 11, GBM 23 and GBM 19. GBM 9 is positive for <i>EGFRvIII</i>. EGFRvIII positive control: U87MGvIII (384 bp), negative control: U87MG (1184 bp) and no template control (NTC): water. B. Sequencing of GBM 9 cDNA confirms the presence of <i>EGFRvIII</i>. C. Real-time amplification plot showing EGFRvIII-positive OSCC patient (13A) and the positive control (U87MGvIII).</p

Specific detection of <i>EGFRvIII</i> mRNA by real-time RT-PCR.

<p>A. Real-time amplification plot of <i>EGFRvIII</i> transcript as detected in the U87MGvIII cell-line. U87MGvIII cells were used as a positive control, U87MG cells were used as negative control: water was used for no template control. B. Direct sequencing of U87MG and U87MGvIII cDNA confirms the presence of <i>EGFRvIII</i> transcript in U87MGvIII cells only. The broken arrow indicates the <i>EGFRvIII</i>-specific exon 1 and exon 8 junction. C. Linear dynamic range of the <i>EGFRvIII</i> real-time RT-PCR assay (100, 10, 1, 0.1 and 0.01 ng cDNA). D. <i>EGFRvIII</i> real-time RT-PCR assay amplification efficiency.</p

Clinicopathological characteristics of OSCC patients evaluated for EGFR protein expression and <i>EGFRvIII</i> frequency.

<p>Clinicopathological characteristics of OSCC patients evaluated for EGFR protein expression and <i>EGFRvIII</i> frequency.</p

CONSORT Diagram outlining patient selection criteria and the different stages at which specific analyses were undertaken for the OSCC study cohort.

<p>CONSORT Diagram outlining patient selection criteria and the different stages at which specific analyses were undertaken for the OSCC study cohort.</p

Immunofluorescent staining and quantitative analysis of EGFR protein expression in OSCC samples using the HistoRx AQUA® platform.

<p>A. Representative examples of normal oral cavity squamous epithelium in the top panel, a low expressing tumor from an <i>EGFRvIII</i>-negative patient in the top-mid panel, and the EGFRvIII negative and EGFRvIIIitive samples from patient 13 in the bottom two panels. DAPI (blue) = nuclei, pan-cytokeratin (green) = epithelial/tumor cells, and EGFR (red). B. Histogram distribution of the EGFR AQUA scores for the entire OSCC patient cohort. The red and green lines indicate the EGFR scores for patient 13 who had both an <i>EGFRvIII</i>-positive (red, Sample ID 13A) and <i>EGFRvIII</i>-negative (green, Sample ID 13B) tumor sample. The solid blue line indicates the median EGFR AQUA score for normal oral cavity squamous epithelium, surrounded by the hashed blue lines representing one standard deviation above and below the median.</p