Cellular internalisation of industrial engineered nanoparticles is undesired and a reason for concern. Here we investigated and compared the ability of seven different mammalian cell cultures in vitro to incorporate six kinds of engineered nanoparticles, focussing on the role of cell type and particle properties in particle uptake. Uptake was examined using light and electron microscopy coupled with energy dispersive X-ray spectroscopy (EDX) for particle element identification. Flow cytometry was applied for semi-quantitative analyses of particle uptake and for exploring the influence on uptake by the phagocytosis inhibitor Cytochalasin D (CytoD). All particles studied were found to enter each kind of cultured cells. Yet, particles were never found within cell nuclei. The presence of the respective particles within the cells was confirmed by EDX. Live-cell imaging revealed the time-dependent process of internalisation of technical nanoparticles, which was exemplified by tungsten carbide particle uptake into the human skin cells, HaCaT. Particles were found to co-localise with lysosomal structures within the cells. The incorporated nanoparticles changed the cellular granularity, as measured by flow cytometry, already after 3 h of exposure in a particle specific manner. By correlating particle properties with flow cytometry data, only the primary particle size was found to be a weakly influential property for particle uptake. CytoD, an inhibitor of actin filaments and therewith of phagocytosis, significantly inhibited the internalisation of particle uptake in only two of the seven investigated cell cultures. Our study, therefore, supports the notion that nanoparticles can enter mammalian cells quickly and easily, irrespective of the phagocytic ability of the cells

The CD4 count is the best surrogate marker for monitoring HIV. The reference method for assessing CD4 counts (flow cytometry, FCM), as expensive technique, is not widely used in non-OECD countries. To make HIV-monitoring available to more patients in these countries, we modified a commercially available density-based negative selection assay (RosetteSep TM ) to make it applicable for low-cost cell enumeration. For evaluation (Step 1), blood taken from 25 HIV patients and 29 healthy donors was assayed with the modified negative selection method (MNS) and compared with FCM. For validation (Step 2), this method was performed in blind quintuplicates on 12 HIVþ blood samples according to FDA guidelines. Association of MNS with the FCM is given by regression models for both steps:Step 1: slope = 1.091, intercept = À46.5.Step 2: slope = 1.074, intercept = À38.3 (Step 2). The imprecision of MNS assessed during Step 2 was 21.2% (intraserial) and 18.8% (interserial). The results suggest that MNS is capable of providing an approximate CD4 count. At a cost of 0.30, it is affordable to patients living in resource-restrained areas. The technique has the potential to deliver an accurate, precise, low-cost test to monitor HIVþ patients. ' 2007 International Society for Analytical Cytology