Going deep: Single cell physiology and cell function

Abstract

function as well as killing assays is of relevance in manyhigh-throughput assays where numerous different effectors,targets, or treatments have to be analyzed. In order to quanti-tate cell function modification by chemical or biologicalagents, several cell-based assays have been developed in thepast. For example, standardized polychromatic (11-color)cytokine expression assays are available to simultaneouslymeasure the intracellular production of five (1) or more cyto-kines in combination with phenotypic markers. Intracellularsignaling patterns can be investigated also, using multiplexedphosphoflow assays (2). In addition, for reliable quantitation,new multiplexed assays are in demand to reduce the numberof assays required, as well as the number of animals needed forreliable results.Inosine triphosphatase (ITPase) polymorphism has beenassociated with several diseases as well as advantageous or dis-advantageous response to therapy. In most studies ITPaseexpression was measured in erythrocyte lysates. However, itwas found that leukocytes express ITPase at substantiallyhigher levels than erythrocytes. Vroemen and coworkers fromHeerlen and Maastricht, The Netherlands (this issue, page672) investigated differences in ITPase levels in different leu-kocyte subsets of human volunteers. To this end, they devel-oped the first polychromatic flow cytometry assays to identifycell phenotypes and quantitate their ITPase expression levels.The authors found that all leukocyte subsets express ITPase.However, expression level was higher in activated monocytesand granulocytes as compared to lymphocytes. These assaysmay help in diseases where ITPase polymorphism plays a rolein order to improve our understanding of disease mechanismsand potentially produce new diagnostic tools for individua-lized therapy.The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide) (MTT) assay is a very commonly used cell-basedvitality assay for high-throughput drug screening. It is based onthe reduction of the yellow water soluble dye MTT into a blueviolet water insoluble dye, formazane (3). Now, Lim and co-workers from the Universities of Seoul and Kongju in Korea(this issue, page 691) developed a microfluidic image cytome-try-based method. Their measurements are multiplexed in thatthey do not only quantitate (on the single cell basis) the meta-bolization of MTT but in addition, also detect cell morphologi-cal changes. This assay was used to test cadmium (Cd