Glioma therapeutic resistance to alkylating chemotherapy is mediated via O6-methylguanine-DNA methyltransferase (MGMT). We hypothesized that a CD45/HAM56/MGMT double-stained cocktail would improve MGMT discrimination in tumor cells versus inflammatory and endothelial cells (IEC). Total MGMT protein was quantified by IHC on 982 glioblastomas (GBM) and 199 anaplastic astrocytomas. Correcting for IEC was done by a CD45/HAM56/MGMT 2-color cocktail. Lowest IEC infiltrates (IEC "cold spots") were identified to quantitate MGMT as well as the percentage of IEC% in the IEC cold spots. MGMT promoter methylation (PM) was also determined. Among the GBM biopsies, mean uncorrected and corrected MGMT% were 19.87 (range 0-90) and 16.67; mean IEC% was 18.65 (range 1-80). Four hundred and fifty one (45.9%) GBM biopsies were positive MGMT PM. Both uncorrected and corrected MGMT% positivity correlated with PM. All 3 MGMT scores correlated with overall survival (OS) in GBM's. Cold spot IEC% was also positively associated with OS. These effects remained in a multivariate model after adjusting for age and disease status. Prognosis determined by correcting MGMT% score for IEC% is not improved in this analysis. However, IEC COLD SPOT score does provide additional prognostic information that can be gained from this correction method.