Data from National Health and Nutrition Examination Survey (NHANES) for 2005-2014 for those aged ≥20 years fasting for ≥8 h (N = 3629) were analyzed to evaluate the role that gender and obesity play in defining correlations between selected perfluoroalkyl substances (PFAS) and total cholesterol (TC), low density lipoprotein cholesterol (LDL), high density lipoprotein cholesterol (HDL), and triglycerides. PFAS considered for analyses were: perfluorooctanoic acid (PFOA), perfluorooctane sulfonate (PFOS), perfluorodecanoic acid (PFDA), perfluorononanoic acid (PFNA), perfluorohexane sulfonate (PFHxS), perfluoroundecanoic acid (PFUnDA), and 2-(N-methyl-perfluorooctane sulfonamido) acetic acid (Me-FOSAA). Gender and obesity stratified regression models were fitted to estimate associations between PFAS and lipid/lipoproteins with adjustments made for confounders. For obese males, but not for nonobese males, positive associations were found between TC and LDL with PFOA (β = 0.0519, p = 0.01 for TC and β = 0.0822, p = 0.03 for LDL), and PFNA (β = 0.0328, p = 0.03 for TC and β = 0.0679, p = 0.04 for LDL). For obese females, adjusted concentrations of TC increased with increase in the concentrations of PFDA (β = 0.0247, p = 0.048), PFNA (β = 0.0286, p = 0.04), and Me-PFOSAA (β = 0.0274, p = 0.02), and there was a positive association of LDL with PFOS (β = 0.0375, p = 0.04), PFDA (β = 0.0397, p = 0.047), and PFNA (β = 0.0593, p = 0.02). The findings, concerning the relationship of longer chain PFAS to serum lipids, suggest greater susceptibility to elevated TC and LDL cholesterol in the obese participants, with some differences between men and women. The key contributing modifiable risk for nonalcoholic steatosis is obesity, and, the development of nonalcoholic steatosis is recognized to be sexually dimorphic. The epidemiologic observation of a susceptible obese subgroup in our data is consistent with toxicology literature findings of disrupted cholesterol metabolism via induced steatosis following PFAS exposure. Gender differences affect serum concentration of PFAS during the reproductive years, and our data add a secondary question concerning whether they also affect the interaction between PFAS exposure and lipid handling in males and females.