Cellometer Vision incorporates image based cell counting and fluorescence detection in a compact and easy-to-use instrument. With fluorescence detection capabilities, Cellometer Vision is an ideal solution for many complex cell population characterization assays such as rapidly counting white-blood-cells in whole blood.
Acridine orange (AO) is a fluorescent nucleic acid stain that has been used to measure nucleated cell concentration in various cell types, including mammalian cell lines, mammalian primary cells, and yeasts. AO is a membrane-permeable dye that stains nucleated cells green.
The following article describes a simple and specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns.
Author institutions: Translational Immunology Section, Office of Science and Technology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, and Translational Autoinflammatory Disease Section, Office of the Clinical Director, National Institute of Arthritis and Musculoskeletal and Skin Diseases
Authors: Barbara Yang, Tuyet-Hang Pham, Raphaela Goldbach-Mansky, Massimo Gadina
Journal: J Vis Exp. 2011; (49): 2662.
A key component of this assay is to standardize the dose response to the number of cells stimulated. They have found 2×106 cells /mL is the sufficient concentration for optimal and measureable concentration of IL-1beta and other cytokines.
“Using Cellometer Vision, diluted blood can be counted without lysing RBC y choosing the whole blood count option using acridine orange (AO). Add one part diluted blood with one part AO and load 20 ml of the 1:1 mixture into a disposable counting chamber. The final AO concentration after dilution should be 1mg/mL. Adjust accordingly with incomplete RPMI 1640 media to final cell centration of 2×106 cells /mL.”
Cellometer Vision has a convenient cell concentration adjustment calculator. Input the original concentration of 20mL, based on the measured cell concentration for each sample, the software calculates the adjustment required to obtain 2×106 cells /mL.