To evaluate the long-term stability of B-type natriuretic peptide (BNP) in frozen samples at −20 °C.During a health and nutrition survey of a representative sample of the adult population of Porto, Portugal, blood samples were collected in EDTA tubes. Aliquots of plasma were frozen at −20 °C until measurement. Subjects with cardiac structural or functional abnormalities or renal dysfunction were excluded. BNP was assessed using a commercial immunofluorescence assay (Triage BNP Test®) as a continuous variable (n=340) and in four subgroups selected according to different storage periods: ≤1 month (n=35), 6–12 months (n=80), 20–24 months (n=18) and ≥36 months (n=21).Age- and sex-adjusted BNP values, analyzed as a continuous variable, decreased significantly over time of storage. When we stratified in four intervals of storage time, the median BNP value (25th–75th percentile, p value) was: 22.6 pg/mL (8.9–76.2, reference group) for ≤1 month, 19.8 pg/mL (9.5–48.4, p=0.055) for 6–12 months, 15.3 pg/mL (7.5–35.0, p=0.037) for 20–24 months, and 2.5 pg/mL (2.5–10.5, p<0.001) for ≥36 months.BNP is stable at −20 °C for 1 year, without protease inhibitors.