Background:Changes occurring in the activity and expression of neutrophils and their transmigration through the blood mammary barrier owing to a mammary infection affecting milk quality and outcome of mastitis.Objective:To understand the role played by various neutrophil molecules on mastitis and milk quality.Methods:18 Karan Fries cows of similar parity, milk yield and lactation stage were selected and screened for mastitis based upon milk Somatic Cell Counts (SCC) as well as California mastitis test and divided into 3 groups of 6 eachi.e., healthy, sub-clinical (SCM), Clinical Mastitis (CM). Milk samples were analyzed for milk composition and technological properties. Milk neutrophils were isolated and their percentage, Phagocytic Activity (PA), viability were estimated. Activities of neutrophil enzymesi.e. Elastase 2, Collagenase and Cathepsin G were analyzed using ELISA. Relative mRNA expression of cell surface molecules like selectin (CD-62L), integrin (CD-11b), chemokine receptors (CXCR1 and CXCR2), CD-44 and chemotactic factor (IL-8) in milk neutrophil were also studied.Results:In the present study, neutrophil percentage (%) was significantly (p˂0.05) higher in SCM and CM milk samples compared to healthy milk samples, whereas neutrophil PA, viability were significantly (p˂0.05) lower in both samples contrast to healthy samples. Activities of Elastase 2 and Collagenase were significantly (p˂0.05) more in SCM milk. There was a significant (p˂0.05) difference in protein, pH, and lactose between healthy, SCM and SCM and CM milk. Significantly (p˂0.05) high Electrical Conductivity (EC) was observed in CM milk than SCM and CM milk. No significant changes in milk fat, Solid Not Fat (SNF) and density were found among any of the groups. The relative mRNA expression of CXCR1, CXCR2 and IL-8 were significantly (p˂0.05) high in milk neutrophils with the progression of SCM and CM, whereas significantly higher expression of CD11b CD-11b was found only in CM cows but there was no change in the expression of CD62L CD-62L in any of the groups. Expression of CD-44 molecule increased significantly in SCM cows, whereas it decreased significantly in clinically infected mastitis cows.Conclusion:This study highlights the changes occurring in the activity of milk neutrophils in healthy, subclinical and clinical mastitis crossbred cows.