Open AccessQuantification of neurotoxicity and identification of cellular subsets in a three‐dimensional brain model
Author(s) -
Pulliam Lynn,
Stubblebine Marcia,
Hyun William
Publication year - 1998
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/(sici)1097-0320(19980501)32:1<66::aid-cyto9>3.0.co;2-d
Subject(s) - acridine orange , ethidium bromide , programmed cell death , confocal microscopy , confocal , cell damage , calcein , microbiology and biotechnology , cell , alexa fluor , biology , chemistry , fluorescence , apoptosis , biochemistry , dna , physics , geometry , mathematics , quantum mechanics , membrane
Imaging of cells in a large intact three‐dimensional tissue remains difficult. Quantification and identification of cell damage in a mixed culture system has been limited by the inability of fluorescent probes to discriminate types of cellular death and penetrate tissue more that 100 μm thick. We have investigated several probes in combination with neural cell‐specific antibodies to quantify cell damage in the presence of several toxins. Acridine orange and ethidium bromide were excellent for determination of cell viability, death by necrosis, or apoptosis in thick brain tissue aggregates. Calcein and ethidium homodimer were effective on live/dead stains, and the Syto dyes 11 and 13 worked well for quantification of all cells in the brain aggregate model. By using these combinations of dyes in conjunction with confocal microscopy, we were able to quantify neural cell damage without disrupting the three‐dimensional environment. Cytometry 32:66–69, 1998. © 1998 Wiley‐Liss, Inc.