To investigate the relationship between acetyl cholinesterase associated collagen gene (COLQ) mutation in patients with acetyl cholinesterase deficiency and its clinical characteristics. Serum and red blood cell acetyl cholinesterase from patients with acetyl cholinesterase deficiency (n=6) and normal controls (n=20) were measured by butyryl thiocholine substrate. COLQ gene variations were detected by sequencing. And the cholinesterase (ChE) genotypes were measured by dibucaine inhibition in vitro. The distributions of ChE surrounded the blood vessels and nerve fibers in lung or pancreas tissues were detected by immunohistochemical staining and indirect immunofluorescence. Serum lactic acid, ammonia and other clinical data were analyzed. Serum ChE in patients with acetyl cholinesterase deficiency were only 1/50 to 1/1000 fold of normal controls. Comparing to controls, dibucaine inhibition values of patients were significantly lower, while there were no differences in red blood cells acetyl cholinesterase. Serum lactic acid and ammonia in patients were significantly higher than controls. Inser 1281-1282 GC of COLQ gene was found in 2 patients, while IVS 6 + 21 T > A, IVS 6 + 30 G > T, IVS 6 + 34 T > C and IVS66 + 12 inser T mutations were found in the other 4 patients, respectively. In addition, the patients with COLQ gene mutation were resistant to regular doses of anesthetics. COLQ gene mutation may be an important reason for the lack of serum ChE in patients with acetyl cholinesterase deficiency.