Quantification of mitogen induced human lymphocyte proliferation: Comparison of alamarblue^tm assay to ³h‐thymidine incorporation assay

Abstract

Proliferation of human lymphocytes in response to various stimuli has traditionally been assessed by measuring uptake of radiolabeled nucleotides such as 3H-thymidine. We have evaluated a fluorometric assay, which uses the commercially available reagent, alamarBlue, as a potential substitute for the 3H-thymidine assay in measuring proliferation of human lymphocytes. In this assay, alamarBlue is added to a population of cells where it is reduced by mitochondrial enzyme activity. The reduced form of the reagent is fluorescent and can be quantitatively detected. The safety and convenience of the alamarBlue assay make it very attractive for use in the clinical laboratory. In this study peripheral blood mononuclear cells (PBMC) from healthy donors were stimulated using the mitogen Concanavalin A, and proliferation was assessed using either the 3H-thymidine or the alamarBlue assay. The alamarBlue assay reliably detects human PBMC and we found that the linear range of detection was 10(4) cells/well (96-well plate) to 5 x 10(5) cells/well. Detection of human PBMC is highly reproducible and the alamarBlue assay may be suitable in a number of applications where detection or relative quantitation of human PBMC is required. The alamarBlue assay also detected mitogen induced proliferation of PBMC although with a significantly lower level of sensitivity than the standard 3H-thymidine assay.

CRA-024781: a novel synthetic inhibitor of histone deacetylase enzymes with antitumor activity in vitro and in vivo

“…Cell Proliferation Assay Ten tumor cell lines and HUVEC were cultured for at least two doubling times, and growth was monitored at the end of compound exposure using an Alamar blue (Biosource, Camarillo, CA) fluorometric cell proliferation assay as previously described (13). The compound was assayed in triplicate wells in 96-well plates at nine concentrations using half-log intervals ranging from 0.0015 to 10 Amol/L.…”

The effect of implemented pulsed electric field (PEF) treatment on the dehydrogenase activity of activated sludge

“…Until now it is not cleared in detail, where the conversion takes place. Zhang et al [17], De Fries and Mitsuhashi [18] assume that it takes place by mitochondrial enzymes in eukaryotic cells. O'Brien et al [19] could only approve at mammalian cells that it takes place in inner site.…”

A new nondestructive cytometric assay based on resazurin metabolism and an organ culture model for the assessment of corneal viability

“…It has been commercialized since 1993 as Alamar blue dye (10) as a viability cell test and has been assessed several times on different types of cells for its cytotoxicity reliability, i.e. fibroblasts (11), immortalized and cancer cell lines (5,12), tumor necrosis factor-hypersensitive cells (13), and for its cell proliferation reliability on mouse and human lymphocytes (6,14) and primary neuronal cell culture (15). Viability measurement using resazurin is similar to or greater than traditional tetrazolium salts (MTT, XTT) and [ 3 H] thymidine assay techniques (5,10,11,16).…”