Method development for the analysis of PBMC-mediated killing of K562 cells by bovine colostrum

Abstract

BackgroundColostrum is the first milk that supplies newborns with immune supporting peptides. Due to its heterogeneous and variable characteristics, standardized assays for assessment of its biological activities are a challenge. The current set of studies were aimed to investigate the immune activity of bovine colostrum blends as well as develop a method to assess variability across different lots. MethodsImmune activity of a bovine colostrum blend was evaluated by their ability to enhance PBMC (peripheral blood mononuclear cell)-mediated killing of K562 cells. K562 cell killing was assessed by flow cytometry using DAPI. PBMCs derived from multiple healthy donors were initially investigated. Frozen PBMC aliquots from one of the highest responders were used for subsequent studies. Different doses and lots of product were assessed. Incubation time was also explored. ResultsBovine colostrum blend similarly reduced K562 cells number and these effects were often greater than the IL-2 positive control. Despite consistent efficacy at enhancing PBMCs-mediated K562 killing, the degree of the effect was significantly variable across different lots. These biological effects were largely dependent on the solubility of the product. ConclusionAssessment of PBMC-mediated killing of K562 cells by flow cytometry using DAPI can be a reliable method for measuring immune activity of bovine colostrum when the material is well-dissolved into solution and the same biological sample from a single donor is used.