TY - JOUR

T1 - Thick methacrylate sections devoid of lost caps simplify stereological quantifications based on the optical fractionator design

AU - Andersen, Stine Hasselholt

AU - Lykkesfeldt, Jens

AU - Larsen, Jytte Overgaard

PY - 2015/9/15

Y1 - 2015/9/15

N2 - AbstractIn neuroscience, the optical fractionator technique is frequently used for unbiased cell number estimations. Although unbiased in theory, the practical application of the technique is often biased by the necessity of introducing a guard zone at one side of the disector to counter lost caps and/or optical limitations. Restricting the disector within the section thickness potentially introduces bias in two ways. Firstly, the need to measure section thickness in order to obtain the disector height/section thickness fraction is challenging since both microcator measurements, microtome block advance, and measurements on re-embedded sections are potentially biased. Secondly, disector placement is not uniform random within the section thickness resulting in a bias in most sections with inhomogeneous cell distribution along the z-axis. Re-embedded 2-hydroxyethylmethacrylate (hereafter methacrylate) sections were inspected for lost caps to evaluate the possibility of whole section thickness counting with the optical fractionator technique and hippocampal granular cell nucleoli density differences along the z-axis were assessed with a z-axis analysis. No lost caps were found in the examined re-embedded tissue and an inhomogeneous cell distribution through the section thickness was observed. In thick methacrylate sections devoid of lost caps sampling through the entire section thickness could be an acceptable alternative to the use of guard zones and the consequent biases associated with section thickness measurement and non-random placement of disectors. This article is protected by copyright. All rights reserved.

AB - AbstractIn neuroscience, the optical fractionator technique is frequently used for unbiased cell number estimations. Although unbiased in theory, the practical application of the technique is often biased by the necessity of introducing a guard zone at one side of the disector to counter lost caps and/or optical limitations. Restricting the disector within the section thickness potentially introduces bias in two ways. Firstly, the need to measure section thickness in order to obtain the disector height/section thickness fraction is challenging since both microcator measurements, microtome block advance, and measurements on re-embedded sections are potentially biased. Secondly, disector placement is not uniform random within the section thickness resulting in a bias in most sections with inhomogeneous cell distribution along the z-axis. Re-embedded 2-hydroxyethylmethacrylate (hereafter methacrylate) sections were inspected for lost caps to evaluate the possibility of whole section thickness counting with the optical fractionator technique and hippocampal granular cell nucleoli density differences along the z-axis were assessed with a z-axis analysis. No lost caps were found in the examined re-embedded tissue and an inhomogeneous cell distribution through the section thickness was observed. In thick methacrylate sections devoid of lost caps sampling through the entire section thickness could be an acceptable alternative to the use of guard zones and the consequent biases associated with section thickness measurement and non-random placement of disectors. This article is protected by copyright. All rights reserved.

KW - Faculty of Health and Medical Sciences

KW - hippocampus

KW - lost caps

KW - methacrylate

KW - stereology

KW - z axis analysis

U2 - 10.1002/ar.23266

DO - 10.1002/ar.23266

M3 - Journal article

C2 - 26370738

VL - 298

SP - 2141

EP - 2150

JO - Anatomical Record

JF - Anatomical Record

SN - 1932-8486

IS - 12

ER -