Introduction: Non-invasive plaque lipid quantification is appealing both for treatment selection and as predictor of future plaque events. However, current MRI methods are insensitive, require a coalesced mass of lipid and rely on multicontrast acquisition. We sought to: (1) provide tissue validation of quantitative T2 maps to measure plaque lipid; and (2) investigate whether this technique could discern differences in plaque characteristics between symptomatic and asymptomatic plaques. We also hypothesised that plaque lipid content correlates with differential plaque macrophage activation. Methods: 40 patients scheduled for carotid endarterectomy were recruited for 3T carotid MRI. Lipid Area was derived from segmented T2 maps and compared directly to plaque lipid defined by histology. Plaque macrophages were procured using laser-capture microdissection from frozen tissue and subjected to microarray analysis. Results: Lipid Area (%) on T2 mapping and histology showed excellent correlation, both by individual slices (R = 0.85, P < 0.001) and plaque average (R = 0.83, P < 0.001). Lipid Area (%) on T2 maps was significantly higher in symptomatic compared to asymptomatic plaques (31.5 ± 3.7% vs. 15.8 ± 3.1%, P = 0.005) despite similar degrees of carotid stenosis. ROC curve analysis showed that T2 mapping has a fair to good ability to discriminate between symptomatic and asymptomatic plaques. Finally, using gene set enrichment analysis, plaque lipid content by T2 map positively correlated with distinct patterns of macrophage activation e.g. INF-STAT1 pathway (P < 0.01, FDR < 0.25). Conclusions: MRI T2 mapping accurately quantifies plaque lipid non-invasively. Greater plaque lipid content was found in symptomatic plaques; and this correlated with inflammatory pathway activation in plaque macrophages. This new technique may find a role in determining optimum treatment e.g. providing an indication for intensive lipid-lowering, or anti-inflammatory therapy.