Subclinical mastitis (SCM) causes huge economic losses in the dairy industry due to suppression in milk production. We have conducted this study to see the infection rate of subclinical mastitis in milking cows of Raninagar Upazila and assess the efficacy of intramammary infusion in cows infected by subclinical mastitis. We have taken milk samples from 103 milking cows and screened for SCM using California Mastitis Test (CMT). We recorded the infection rate 51.5 % for SCM and treated the udder with the udder infusion of Mastanil® drug containing Gentamicin sulphate 100mg/ml supplied by Techno Drugs Ltd, Bangladesh. The infection rate was remarkably increased with the age and parity of cows. The prevalence of 33.1% was notably found in the left hind (LH) quarter compared to other quarters. We have also found that the effectiveness of intramammary infusion with Mastanil® drug in lactating cows is 50.9%. Despite limitations regarding accurate disease investigation procedure, we conducted our experiments, thus, further research is warranted to observe the hygiene regarding milker’s hand and milking instruments and dry cow therapy with other potential drug to assess and validate our observations in managing SCM substantially.