Eaton S.L.,Roslin Institute |
Cumyn E.,Roslin Institute |
Cumyn E.,University of Edinburgh |
King D.,Roslin Institute |
And 6 more authors.
Journal of Anatomy | Year: 2016
Quantification of immunohistochemically (IHC) labelled tissue sections typically yields semi-quantitative results. Visualising infrared (IR) 'tags', with an appropriate scanner, provides an alternative system where the linear nature of the IR fluorophore emittance enables realistic quantitative fluorescence IHC (QFIHC). Importantly, this new technology enables entire tissue sections to be scanned, allowing accurate area and protein abundance measurements to be calculated from rapidly acquired images. Here, some of the potential benefits of using IR-based tissue imaging are examined, and the following are demonstrated. Firstly, image capture and analysis using IR-based scanning technology yields comparable area-based quantification to those obtained from a modern high-resolution digital slide scanner. Secondly, IR-based dual target visualisation and expression-based quantification is rapid and simple. Thirdly, IR-based relative protein abundance QIHC measurements are an accurate reflection of tissue sample protein abundance, as demonstrated by comparison with quantitative fluorescent Western blotting data. In summary, it is proposed that IR-based QFIHC provides an alternative method of rapid whole-tissue section low-resolution imaging for the production of reliable and accurate quantitative data. © 2016 Anatomical Society.
Devlin A.-C.,University of St. Andrews |
Devlin A.-C.,Euan MacDonald Center for Motor Neurone Disease Research |
Burr K.,Euan MacDonald Center for Motor Neurone Disease Research |
Burr K.,University of Edinburgh |
And 12 more authors.
Nature Communications | Year: 2015
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease for which a greater understanding of early disease mechanisms is needed to reveal novel therapeutic targets. We report the use of human induced pluripotent stem cell (iPSC)-derived motoneurons (MNs) to study the pathophysiology of ALS. We demonstrate that MNs derived from iPSCs obtained from healthy individuals or patients harbouring TARDBP or C9ORF72 ALS-causing mutations are able to develop appropriate physiological properties. However, patient iPSC-derived MNs, independent of genotype, display an initial hyperexcitability followed by progressive loss of action potential output and synaptic activity. This loss of functional output reflects a progressive decrease in voltage-activated Na + and K + currents, which occurs in the absenceof overt changes in cell viability. These data implicate early dysfunction or loss of ion channels as a convergent point that may contribute to the initiation of downstream degenerative pathways that ultimately lead to MN loss in ALS. © 2015 Macmillan Publishers Limited.