The current study was designed to examine the association between a composite Healthy Lifestyle Score (HLS) and thyroid function biomarkers among American adults. This cross-sectional study utilized data from 5,693 adults aged ≥18 years in the NHANES 2007–2012 cycles. A HLS (range 0–6) was constructed based on six modifiable factors: non-smoking, no heavy alcohol intake, normal BMI (18.5–24.9 kg/m²), high physical activity (upper tertile of MET-min/week), adequate sleep (7–9 h/night), and appropriate energy intake. Serum concentrations of thyroid-stimulating hormone (TSH), free and total thyroxine (FT4, TT4), free and total triiodothyronine (FT3, TT3), thyroglobulin (Tg), and thyroid antibodies (TPOAb, TgAb) were measured. Multivariable linear regression adjusted for sociodemographic factors was used to assess associations. In fully adjusted models, each one-point increase in HLS was associated with lower serum FT4 (β = −0.07 ng/dL; 95% CI: −0.10, −0.03; p < 0.001) and TT4 (β = −0.11 µg/dL; 95% CI: −0.15, −0.06; p < 0.001). Compared with participants with an HLS of 0–1, those with HLS 4–6 had lower FT4 (β = −0.20; 95% CI: −0.30, −0.09; p < 0.001) and TT4 (β = −0.36; 95% CI: −0.49, −0.22; p < 0.001). Associations for other thyroid markers were not statistically significant after correction for multiple comparisons (p > 0.05). A healthier lifestyle is inversely associated with serum FT4 and TT4 levels, highlighting potential links between modifiable behaviors and thyroid physiology.