Foi comparada a efetividade da anti-sepsia de sítios operatórios em vinte e quatro animais, subdivididos em três grupos, utilizando água destilada (grupo controle), álcool-iodo-álcool (grupo I) e gluconato de clorexidina (grupo II). As amostras foram coletadas através de swab da pele, depois da tricotomia (T0), após anti-sepsia (T1) e duas horas após o uso do anti-séptico (T2), e submetidas à contagem de Unidades Formadoras de Colônia(UFC)/ml. Nos três grupos, ocorreu crescimento bacteriano em T0; no T1 a redução média de UFC/ml foi de 26,70% para o grupo controle, 91,61% para o grupo I e 96,67% para o grupo II. No T2, as reduções nos respectivos grupos foram de 21,02%, 91,56% e 96,89%. As duas técnicas utilizando anti-sépticos reduziram significativamente o número de bactérias da pele, tanto no T1 quanto no T2 (pThe efficacy of the antisepsis in a surgical area was compared in twenty four mongrel dogs, divided in three groups, using sterile water, alcohol-iodine-alcohol and chlorhexidine gluconate. All samples of skin swab were collected after thricothomy, after antisepsis, and two hours after application of the antiseptic solution. Samples were collected with a sterile swab and the Colony Formation Unities/ml was determined. Bacterial growth were observed in samples from all groups during the first collection. However, at the first counting (T1), the average reduction of CFU/ml observed was 26.7% for the control group, 96.61% group one, and 96.67% group two. At the second counting (T2), reductions of 21.02%, 91.56%, and 96.89% for the respective groups were observed. The two antiseptic techniques utilized were able to reduced significantly the number of bacteria present on the skin, at T1 as well as T2 (p<0.05). The equality of these two antiseptic methods was demonstrated since no significant difference (p<0.05) between these treatments was observed.