An experimental flow cytometer was constructed using a quartz flow cell optically coupled to a 1.22 NA lens. A pair of crossed cylindrical quartz lenses allowed multilaser excitation. Two helium-cadmium (HeCd) lasers, emitting 16 mW at 442 nm and 35 mW at 325 nm, were used to excite chromomycin A3 and Hoechst 33258 fluorescence, respectively. Bivariate flow karyotypes from normal human male chromosomes and from the Daudi cell line were obtained and were compared to those from a standard instrument using dual water-cooled lasers. The new experimental instrument exhibited comparable resolution to that from the standard instrument. In further experiments with Daudi chromosomes, the 35 mW HeCd laser was replaced with a 10 mW HeCd laser, and the system still gave good, though slightly decreased, resolution.

We have previously shown that inhaled Con A has a powerful enhancing effect on the formation of immune complexes between an inhaled antigen and circulating antibody. Immunohistochemical staining has demonstrated such complexes, together with host complement, in close association with foci of necrotizing destruction of the pulmonary parenchyma. We have postulated that Con A promotes immune complex formation indirectly through polyclonal activation of lymphocytes in the lung. In this paper we test this hypothesis in animals rendered unresponsive to Con A stimulation in vivo by i.v. administration of cholera toxin (CT). Such treatment raised the levels of cAMP in peripheral blood lymphocytes and inhibited their proliferative response to Con A in vitro. CT administration further blocked the local inflammatory response to intradermal injections of Con A, as well as the cell-mediated immune response to intradermal injections of BSA. Although CT failed to block the immune complex-mediated Arthus vasculitis in the skin, it did block production of immune complex pulmonary injury by antigen/mitogen aerosols, as did decomplementation with purified cobra venom factor. These findings support the hypothesis that polyclonal activation of pulmonary lymphocytes promotes immune complex-type alveolitis, possibly by facilitating interactions between humoral antibody and intra-alveolar antigen.

A dextran density gradient centrifugation method has been used to separate PHA transformed lymphocytes from nontransformed lymphocytes obtained from rat blood. Fifty-four percent of the transformed lymphocytes were recovered in the fraction at a density of 1.0620 g/ml. Eighty-five percent of the cells in this fraction were transformed cells while the remainder were non-transformed lymphocytes.