, 1983a, Boyer et al., 1983b, Gibbs et al., 1994, Moore et al., 2003 and Law selleck products et al., 2005), but has not eradicated GBS disease in infants ( Schuchat, 2000 and Phares et al., 2008). GBS is still a public health concern and the introduction of additional prevention and therapeutic strategies is highly desirable. During the last two decades, polysaccharide-protein conjugate vaccines against GBS have been extensively

studied in preclinical and human clinical studies ( Baker et al., 1999, Baker et al., 2000, Baker et al., 2003a, Baker et al., 2003b, Baker et al., 2007, Lancaster et al., 2011 and Heath, 2011). An obstacle to the development of vaccines against GBS is the difficulty of conducting clinical efficacy trials, because of the relatively low incidence of neonatal GBS disease. A possible solution to overcome this difficulty may come from the development of an effective functional antibody assay as in vitro correlate of protection. The most commonly used method to assess functional antibodies to GBS in post-immunization sera is the in vitro killing-based opsonophagocytosis assay (kOPA) that mimics the in vivo process of killing by host effector

cells, following opsonization by specific antibodies ( Baltimore et al., 1977, Edwards et al., 1979 and Guttormsen et al., 2008). This assay can constitute a viable surrogate of the effectiveness check details of a GBS vaccine, as passive protection of mice by sera from individuals immunized with GBS polysaccharide-based vaccines correlated with high functional antibody titers measured by OPA ( Baltimore et al., 1981, Kasper et al., 1996 and Brigtsen et al., 2002). However, bacterial growth, colony plating and counting are time and resource consuming MycoClean Mycoplasma Removal Kit steps and standardization presents challenges due to the source and quality of effector cells and to the variability associated with plating and colony counting.

Although cultured phagocytes (differentiated HL-60 cells) can be used in place of human peripheral polymorphonuclear leukocytes (PMNs), as a less variable neutrophil source ( Romero-Steiner et al., 1997 and Guttormsen et al., 2008,), the fraction of HL-60 cells differentiated to active phagocytes varies, representing a further source of variability. Fluorescence based OPAs can limit the effort and variability associated with plating and counting of surviving bacteria (Plested and Coull, 2003, Guttormsen et al., 2009 and Simons, 2010,). These assays use bacteria labeled with fluorophores, such as fluorescein-derivatives (dicarboxyfluorescein, dihydrodichlorofluorescein, Oregon green), rhodamine or Alexa Fluor derivatives (Rodríguez et al., 2001 and Guttormsen et al., 2009) and are rapid and efficient for large scale testing of sera. However, these approaches do not distinguish between adherent and internalized bacteria.