Little is known regarding factors associated with soluble amyloid beta peptide (Aβ) concentrations in humans at late midlife, when Aβ is likely most critical to Alzheimer disease pathogenesis. We examined the association between insulin, insulin-related factors, and plasma Aβ at late midlife. Plasma Aβ42, Aβ40, fasting insulin, and c-peptide were measured in 468 women without diabetes, aged 59 to 69 years (median 63 y). Before blood draw, participants reported body mass index, waist circumference, physical activity, alcohol intake, hypertension, and diabetes family history. Linear regression was used to calculate age-adjusted mean differences in Aβ42 to Aβ40 ratio, and Aβ42 levels, by insulin and insulin-related factors. The ratio of Aβ42 to Aβ40 was statistically significantly lower in women with family history of diabetes, and Aβ42 was significantly lower with less physical activity, greater waist circumference, hypertension, and family history of diabetes (P<0.05 for all). Aβ42 to Aβ40 ratio, and Aβ42 levels, appeared lower with higher c-peptide levels (P trend=0.07 and 0.06, respectively), although these were not statistically significant. In summary, insulin-related factors appear associated with lower plasma Aβ42 to Aβ40 ratio, and Aβ42, at late midlife, consistent with increased brain sequestration of Aβ42 (relative to Aβ40), suggesting insulin merits focus in strategies to prevent dementia.