Microscopic differential cell counting and viability evaluation of milk somatic cells during lactation in goat

Milk contains different cell populations commonly known as somatic cells, changing their distribution under the influence of several factors as physiological status and stage of lactation, and useful to assess the transcriptional activity of the mammary gland in livestock. The aim of this study was to compare the differential somatic cells counting (DCCs) and cell viability in goat milk samples collected at morning milking (MM) and 2 hours later (2H), in order to verify if some difference occurred between the two sampling times. This investigation was carried out within a project addressed to optimize studies on gene expression in goats mammary gland. In 11 pluriparous Alpine goats lactations were studied for 21 wks. Individual milk yield was recorded in three different time points (TP), at 38±5 days (TP0), 91±7 days (TP1) and 140±7 days (TP2) of lactation. Milk samples were collected twice a day: at MM and 2H, in the different TPs. For DCCs, 150 mL of milk were centrifuged and the pellet obtained was washed and resuspended in PBS. Slides were smeared and stained with May Grunwald-Giemsa stain. Two hundred cells were evaluated per slide under a light microscope, classified as neutrophils, lymphocytes, macrophages and epithelial cells, and expressed as a percentage of the total number of cells counted. Cell viability was estimated by trypan blue exclusion test. Two hundred cells were counted using a Neubauer's chamber, and discriminated as live or dead. A two-way analysis of variance (General Linear Model, PROC GLM, SAS Institute) was used to analyse the effects of TP and milking distance (MM and 2H) on milk somatic cells composition and viability. Treatment effects were declared significant at P<=0.05. Percentage of neutrophils increased significantly during lactation, whereas % of lymphocytes, macrophages and epithelial cells decreased. The percentage of viable cells in milk increased significantly from MM to 2H samples (63.97±2.35 and 77.01±2.25, respectively) but was not affected by lactation stage (TP0=70.0±3.4; TP1=69.5±2.4; TP2=72.0±2.4). Moreover the milking distance didn't affect somatic cells composition (neutrophils: MM=54.7±2.9, 2H=55.6±3.5; lymphocytes: MM=35.4±2.5, 2H=33.4±3.0; macrophages: MM=3.9±0.3, 2H=4.1±0.4; epithelial cells: MM=5.7±0.8, 2H=6.6±1.0). Results obtained in this study suggest that somatic cells are more viable and transcriptionally active in samples collected 2 hours after morning milking.