The accumulation and aggregation of the beta‐amyloid peptide (Aβ) in the brain represents a key factor in the pathogenesis of Alzheimer's disease (AD). Many of the transgenic mouse models for AD exhibit an amyloid pathology with neuritic plaques but they typically vary by the type and abundance of plaques identified in their brains and by the onset and severity of cognitive impairment. Thus, an important consideration in the characterization of AD transgenic mouse models should be the quantitative evaluation of the amyloid load in the brain together with a detailed physico‐chemical analysis of Aβ from the deposited plaques. Here we present an analytical procedure to collect single amyloid plaques from anatomically defined brain regions by laser dissection microscopy that can be quantitatively assessed in their Aβ isoforms composition by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry. Quantification was achieved by stable isotope dilution using calibrated ¹⁵N‐labeled Aβ standards that were spiked in the sample immediately after laser dissection. Using this method, we found that the amyloid loads in brain plaques isolated from the transgenic AD mouse model PS2APP or from human were similar. Total Aβ composition was estimated at ∼50–100 fmol per excised plaque disc, as confirmed by immunoblot analysis. N‐Terminal truncated Aβ isoforms were identified in both transgene and human amyloid plaques but with significantly elevated levels in human samples. Copyright © 2005 John Wiley & Sons, Ltd.