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The aim of the study was to examine the effects of promoting increased lean red meat consumption on serum concentrations of total and high-density lipoprotein (HDL) cholesterol, and serum fatty acid composition, among toddlers. In a 20-week randomized controlled trial healthy 12 to 20-month-old children were assigned to: red meat (n = 90; parents were encouraged to add 56 g/day of lean red meat to their toddler's usual diet), or control (n = 90) groups. Food and nutrient intakes were assessed with 3-day weighed food records (baseline, week 4, and week 20). Serum was analyzed for total and HDL cholesterol concentrations, and fatty acid composition (baseline and week 20). At week 20, relative to control, the red meat group had higher intakes of red meat, all meat, and carbohydrate; and lower intakes of milk, energy, cholesterol, and total, saturated, and monounsaturated fat (P = 0.043 for energy, all others P ≤ 0.002). No effects associated with the intervention were found for total cholesterol, HDL cholesterol, total to HDL cholesterol ratio, or serum fatty acid composition (all P ≥ 0.059) aside from pentadecanoic acid (P = 0.047). An ∼3-fold increase in lean red meat intake, from ∼10 to ∼30 g/day, resulted in no consistent changes in serum lipids or fatty acid composition, suggesting that the addition of ∼2 tablespoons/day of lean red meat to toddlers’ diets will likely not adversely affect serum lipids or serum fatty acids.