Insulin and insulin-like growth factor (IGF)-1 signaling is a proposed mechanism linking dietary protein and major chronic diseases. However, it is unclear whether animal and plant proteins are associated with biomarkers of insulin and IGF axis. We analyzed a total of 14,709 participants from Nurses’ Health Study and Health Professionals Follow-up Study who had provided a blood sample. Detailed dietary information was assessed using validated food frequency questionnaires. We assessed C-peptide, insulin, IGF-1, and IGF binding proteins (BP). Multivariable-adjusted linear regressions were used to examine associations of animal and plant protein intake with biomarkers after adjusting for confounders. The medians (5th-95th percentiles) of animal and plant protein intake (% of total energy) were 13% (8–19%) and 5% (4–7%), respectively. Compared to participants in the lowest quintile, those in the highest quintile of animal protein had 4.8% (95% CI: 1.9, 7.9; P-trend<0.001) higher concentration of IGF-1 and -7.2% (95% CI: −14.8, 1.1; P for trend = 0.03) and −11.8% (95% CI: −20.6, −1.9; P-trend<0.001) lower concentration of IGFBP-1 and IGFBP-2, respectively, after adjustment for major lifestyle factors and diet quality. In contrast, no association was observed between animal protein intake and C-peptide, insulin and IGFBP-3. The associations were restricted to participants with at least one unhealthy lifestyle risk factor (i.e., overweight/obese, physical inactivity, smoking, and heavy alcohol intake). Plant protein tended to be strongly associated with numerous biomarkers in age-adjusted analyses but these became largely attenuated or non-significant in multivariable adjustment. Plant protein intake remained positively associated with IGF-1 (P-trend = 0.002) and possibly IGFBP-1 (P-trend = 0.02) after multivariable adjustment. Substitution of plant protein with animal protein sources was associated with lower IGFBP-1. In additional analysis, IGF-1 and IGFBPs were estimated to mediate approximately 5–20% of the association between animal protein and type 2 diabetes. Higher animal protein intake was associated with higher IGF-1 and lower IGFBP-1 and IGFBP-2, whereas higher plant protein intake was associated with higher IGF-1 and IGFBP-1.