Abstract
The large size of primate brains is an impediment to obtaining high-resolution cell number maps of the cortex in humans and non-human primates. We present a rapid, flow cytometry-based cell counting method that can be used to estimate cell numbers from homogenized brain tissue samples comprising the entire cortical sheet. The new method, called the flow fractionator, is based on the isotropic fractionator (IF) method (Herculano-Houzel and Lent, 2005), but substitutes flow cytometry analysis for manual, microscope analysis using a Neubauer counting chamber. We show that our flow cytometry-based method for total cell estimation in homogenized brain tissue provides comparable data to that obtained using a counting chamber on a microscope. The advantages of the flow fractionator over existing methods are improved precision of cell number estimates and improved speed of analysis.
Highlights
Cell numbers vary across areas and regions of the cerebral cortex in primates, there are currently only low-resolution, incomplete or no cell number maps available for some primate species, including humans (Collins et al, 2010b)
We present a new method, the flow fractionator, that is based on the isotropic fractionator (IF) method, but uses flow cytometry to count the total numbers of cells in a homogenized suspension, rather than manual counting on a fluorescence microscope
The flow fractionator method resulted in repeatable cell population estimates that were in excellent agreement with data collected on the microscope using the IF method
Summary
Cell numbers vary across areas and regions of the cerebral cortex in primates, there are currently only low-resolution, incomplete or no cell number maps available for some primate species, including humans (Collins et al, 2010b). Highresolution cell number maps, in large-brained primate species like the human will require cell counts in thousands of small cortical samples to more fully characterize the total cell numbers in identifiable cortical areas and regions and overall patterns of cell distribution (Collins, 2011). Stereological methods can be used to estimate the total number of cells, their shape, size, and volume in sectioned and stained tissue (e.g., Schmitz and Hof, 2005). These methods are best suited to cell population estimates of small, homogeneous brain areas with well-defined boundaries and are less practical for producing detailed cell number maps across the whole cortex from large brains. Using manual counting on a Neubauer counting chamber and a fluorescence microscope, the number of nuclei is estimated in a small aliquot from the nuclear suspension, and the data are extrapolated to the total suspension volume using the published formula
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
