Correlating short-term Ca(2+) responses with long-term protein expression after activation of single T cells.
In order to elucidate the dynamics of cellular processes that are induced in context with intercellular communication, defined events along the signal transduction cascade and subsequent activation steps have to be analyzed on the level of individual cells and correlated with each other. Here we present an approach that allows the initiation of cell-cell or cell-particle interactions and the analysis of cellular reactions within various regimes while the identity of each individual cell is preserved. It utilizes dielectrophoresis (DEP) and microfluidics in a lab-on-chip system. With high spatial and temporal precision we contacted single T cells with functionalized microbeads and monitored their immediate cytosolic Ca(2+) response. After this, the cells were released from the chip system and cultivated further. Expression of the activation marker molecule CD69 was analyzed the next day and correlated with the previously recorded Ca(2+) signal for each individual cell. We found a significant difference in the patterns of Ca(2+) traces between activated and non-activated cells, which shows that Ca(2+) signals in T cells can provide early information about a later reaction of the cell. Although T cells are non-excitable cells, we also observed irregular Ca(2+) transients upon exposure to the DEP field only. These Ca(2+) signals depended on exposure time, electric field strength and field frequency. By minimizing their occurrence rate, we could identify experimental conditions that caused the least interference with the physiology of the cell.
Kirschbaum
Michael
M
Fraunhofer Institute for Biomedical Engineering (IBMT), Am Muehlenberg 13, 14476 Potsdam, Germany.
Jaeger
Magnus Sebastian
MS
Duschl
Claus
C
D000906:Antibodies / Q000276:immunology; D015703:Antigens, CD / Q000378:metabolism*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000378:metabolism*; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000276:immunology; D002455:Cell Division / Q000276:immunology; D002470:Cell Survival / Q000276:immunology; D003600:Cytosol / Q000378:metabolism; D004586:Electrophoresis; D005786:Gene Expression Regulation; D006801:Humans; D019169:Jurkat Cells; D037181:Lectins, C-Type; D008213:Lymphocyte Activation; D008564:Membrane Potentials / Q000276:immunology; D046210:Microfluidic Analytical Techniques; D008863:Microspheres; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D013997:Time Factors; D000099192:CD69 Antigens
D000906:Antibodies; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D018106:CD28 Antigens; D017252:CD3 Complex; D002118:Calcium; D037181:Lectins, C-Type; D000099192:CD69 Antigens