The study was designed to assess the effects of in vitro selenium addition on intracellular hydrogen peroxide production by neutrophils from the milk and blood of dairy cows. Blood from 10 dairy cows and 20 milk samples from ϐive dairy cows were incubated with 0 mg (control) or 10μM of sodium selenite. Then, milk and blood neutrophils were submitted for evaluation of intracellular hydrogen peroxide production by ϐlow cytometry using 2',7'-dichloroϐluorescein diacetate as a probe. The selenium status of the animals was evaluated by determination of the blood glutathione peroxidase activity. The results of the present work showed that in vitro selenium supplementation leads to an enhancement in intracellular hydrogen peroxide production, which indicates an improvement in the bactericidal effects of blood and milk neutrophils even in cows with a selenium-adequate status. Thus, the present study showed that in vitro Se supplementation leads to an enhancement in intracellular hydrogen peroxide production, indicating an improvement in the bactericidal effects of blood and milk neutrophils in cows with Se-adequate status. A determinação da produção intracelular de peróxido de hidrogênio se deu por citometria de ϐluxo através da utilização do 2´,7´ diclorodihidroϐluoresceína diacetato como sonda. A mensuração do conteúdo de selênio foi avaliada pela atividade da glutationa peroxidase eritrocitária. Os leucócitos polimorfonucleares tanto sanguíneos quanto do leite apresentaram signiϐicativo aumento na produção intracelular de H 2 O 2 com a suplementação in vitro de selênio. Desta forma, o presente estudo apontou para aumento da produção intracelular de H 2 O 2 , indicando aumento da capacidade microbicida dos leucócitos polimorfonucleares sanguíneos e lácteos mesmo em animais com níveis adequados de selênio.