%0 Journal Article
%T A novel flow cytometric assay for measurement of In Vivo pulmonary neutrophil phagocytosis
%A Elizabeth A Vander Top
%A Greg A Perry
%A Martha J Gentry-Nielsen
%J BMC Microbiology
%D 2006
%I BioMed Central
%R 10.1186/1471-2180-6-61
%X The viability of the alveolar macrophages and PMNs isolated from the lavage fluid was >95%. The values of the percentage of PMNs in the lavage fluid as well as the percentage of PMNs associated with CFSE-labeled S. pneumoniae as measured through flow cytometry showed a high degree of correlation with the results from manual counting of cytospin slides.This assay is suitable for measuring bacterial uptake within the infected lung. It can be adapted for use with other organisms and/or animal model systems.Phagocytosis and killing of pathogens by resident alveolar macrophages and recruited polymorphonuclear leuckocytes (PMNs) is integral in clearing bacterial respiratory tract infections. Because alveolar macrophages (AM) do not efficiently phagocytose most strains of Streptococcus pneumoniae (the pneumococcus), uptake and killing by recruited PMNs is particularly important during pneumococcal pneumonia [1]. For a number of years, our laboratory has been studying the effects of ethanol ingestion and liver disease on pulmonary PMN function. We began with traditional phagocytosis assays performed in vitro with peripheral or peritoneal PMNs and heat-killed, pre-opsonized organisms. Such assays are limited, however, in that they cannot adequately mimic the conditions present within the infected lung. Therefore, they may not detect all host defects leading to reduced phagocytic uptake of bacteria during pulmonary infections.One particular difficulty with the use of in vitro assays for measurement of pneumococcal phagocytosis is that the organisms are not taken up unless they are pre-opsonized with serum containing active complement components, even though the serum may contain high titers of specific antipneumococcal antibodies [2]. This need for pre-opsonization of the organisms makes it impossible to examine certain underlying host immune defects such as complement deficiencies. The serotype 3 pneumococcal strain used in our studies (ATCC 6303) presents an additional uniq
%U http://www.biomedcentral.com/1471-2180/6/61