In this issue

Membrane microdomains are selectively “helpful”

Polarized helper T lymphocytes are key players in orchestration of immune responses through their distinct cytokine production and secretion. Using various flow and image cytometric methods, Izsepi and coworkers studied the spatial organization of T-cell receptors, the related signaling molecules and lipid rafts in model hybridomas of Th0, Th1 and Th2 phenotype, as well as in primary splenic T cells polarized in appropriate cytokine milieu. Their data firmly support that differential expression and spatial distribution of several lipid and protein constituents of T-cell signaling rafts at the cell surface may define the nature of the subsequent calcium signal and through this the nuclear residence time of NFAT transcription factor. The latter is of key importance in activation of cytokine genes and thus in the polarized T-cell effector response to antigens. The analysis presented here may also help to designate signal therapeutic targets in treatment of diseases linked to unbalanced Th1 or Th2 cell activation. 1

Illustration 1.

In this issue: page 185

How accurate can cell viability be measured?

Metrological research in the area of health care supports efficient use of diagnostic techniques. Quantification of cell viability is a demanding yet important task for cell research, e.g. for regenerative medicine. Different measurement techniques, staining protocols and data analysis methods lead to methodical influences which may significantly contribute to the uncertainty of measurement. Kummrow and coworkers addressed this issue by comparing flow cytometry and microscopy, two commonly used methods to assess cell viability. In each case, fluorescence was detected utilizing live/dead staining by calcein acetoxymethyl-ester and ethidium homodimer-1. Peripheral blood monocytes and circulating stem cells were used as a model to assure an adequate comparison for both modalities in a wide range of cell viabilities. On average, microscopy and flow cytometry yield equal results provided that sufficiently high numbers of cells are counted. Although specific applications may favor one of the investigated methods, both of them complement each other. 2

Illustration 2.

In this issue: page 197

A Universal Nanoparticle Cell Secretion Capture Assay

Secreted proteins play an important role in intercellular interactions. Fitzgerald and Grivel have developed a universal assay that allows a simple non-invasive identification and isolation of cells based on their secretion of various products. Their method is based on the targeting, to the cell surface, of hetero-functional nanoparticles coupled to a cell-surface specific antibody and to a secreted-protein-specific antibody, which captures the secreted protein on the surface of the secreting cell. This method allows virtually limitless flexibility and furthermore does not compromise cell viability allowing culture and expansion of the secreting cells. 3

Illustration 3.

In this issue, page 205

Discover New Drugs with Diverse Efficacies, Fast

Cell surface receptors such as G-protein coupled receptors, receptors for tyrosine kinases, and ion channels are major drug targets. Ligand induced trafficking of these receptors can serve as a useful therapeutic model. However, direct measurement of plasma membrane protein trafficking by flow cytometry has been challenging. Wu and coworkers have developed a hybrid platform which allows drug discovery for trafficking receptors, previously not suitable for flow cytometry or high throughput screening, using the recently available reporter protein tag based on the fluorogen activating protein. The robust, sensitive and highly versatile platform has the potential to identify both canonical and non-canonical ligands of target receptors, and to investigate the ligand efficacies associated with the receptor internalization pathway. The system has been extended to simultaneously monitor the behavior of two different receptors in a single well, which opens the door to the investigation of receptor/co-receptor pairs arising from synergistic drug effects. 4

Illustration 4.

In this issue, page 220