Kidney disease is common among captive cheetahs (Acinonyx jubatus). Serum creatinine is the most common measurement to estimate glomerular filtration rate (GFR) because of the ease of its clinical use, but it is a crude estimate that only increases after significant disease is already present and is affected by extrarenal factors. Symmetric dimethylarginine (SDMA) is a renal biomarker in humans, dogs, and cats that correlates with serum creatinine and GFR and appears to be an earlier and more specific biomarker for kidney disease. Ninety-two banked serum samples from 11 cheetahs housed at the Oklahoma City Zoo from 1992 to 2012 were retrospectively analyzed. Histopathology results were available for 10/11 cheetahs, and all 10 had histologic renal lesions. General categories of renal lesions included glomerulosclerosis (7/10; 70%), amyloidosis (7/10; 70%), inflammatory (9/10; 90%), and oxalate nephrosis (2/10; 20%). SDMA immunoassay and mass spectrometry were measured for validation and compared with creatinine to assess for correlation. Serum creatinine concentrations were determined by enzymatic colorimetric methods. SDMA immunoassay was validated in cheetahs and correlated well with serum creatinine (R²=0.687; P < 0.0001). SDMA and serum creatinine measured from freeze-thawed stored samples show high correlation in individual cheetahs (R² = 0.972; P < 0.0001). These data support that SDMA could be a promising renal biomarker in cheetahs. Further research is warranted to investigate whether SDMA might be an earlier indicator of kidney disease in cheetahs and whether this assay can be extended to other nondomestic carnivores.