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Separation of photoreceptor cell compartments in mouse retina for protein analysis

Overview of attention for article published in Molecular Neurodegeneration, April 2017
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Title
Separation of photoreceptor cell compartments in mouse retina for protein analysis
Published in
Molecular Neurodegeneration, April 2017
DOI 10.1186/s13024-017-0171-2
Pubmed ID
Authors

Kasey Rose, Steven T. Walston, Jeannie Chen

Abstract

Light exposure triggers movement of certain signaling proteins within the cellular compartments of the highly polarized rod photoreceptor cell. This redistribution of proteins between the inner and outer segment compartments affects the performance and physiology of the rod cell. In addition, newly synthesized phototransduction proteins traverse from the site of their synthesis in the inner segment, through the thin connecting cilium, to reach their destination in the outer segment. Processes that impede normal trafficking of these abundant proteins lead to cell death. The study of movement and unique localization of biomolecules within the different compartments of the rod cell would be greatly facilitated by techniques that reliably separate these compartments. Ideally, these methods can be applied to the mouse retina due to the widespread usage of transgenic mouse models in the investigation of basic visual processes and disease mechanisms that affect vision. Although the retina is organized in distinct layers, the small and highly curved mouse retina makes physical separation of retinal layers a challenge. We introduce two peeling methods that efficiently and reliably isolate the rod outer segment and other cell compartments for Western blots to examine protein movement across these compartments. The first separation method employs Whatman(®) filter paper to successively remove the rod outer segments from isolated, live mouse retinas. The second method utilizes Scotch(TM) tape to peel the rod outer segment layer and the rod inner segment layer from lyophilized mouse retinas. Both procedures can be completed within one hour. We utilize these two protocols on dark-adapted and light-exposed retinas of C57BL/6 mice and subject the isolated tissue layers to Western blots to demonstrate their effectiveness in detecting light-induced translocation of transducin (GNAT1) and rod arrestin (ARR1). Furthermore, we provide evidence that RGS9 does not undergo light-induced translocation. These results demonstrate the effectiveness of the two different peeling protocols for the separation of the layered compartments of the mouse retina and their utility for investigations of protein compositions within these compartments.

Twitter Demographics

The data shown below were collected from the profiles of 2 tweeters who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 13 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Slovenia 1 8%
Unknown 12 92%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 4 31%
Student > Postgraduate 3 23%
Researcher 3 23%
Professor > Associate Professor 1 8%
Other 1 8%
Other 0 0%
Unknown 1 8%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 4 31%
Agricultural and Biological Sciences 4 31%
Neuroscience 3 23%
Sports and Recreations 1 8%
Unknown 1 8%

Attention Score in Context

This research output has an Altmetric Attention Score of 1. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 13 April 2017.
All research outputs
#5,464,984
of 9,684,470 outputs
Outputs from Molecular Neurodegeneration
#356
of 481 outputs
Outputs of similar age
#149,008
of 263,642 outputs
Outputs of similar age from Molecular Neurodegeneration
#17
of 19 outputs
Altmetric has tracked 9,684,470 research outputs across all sources so far. This one is in the 26th percentile – i.e., 26% of other outputs scored the same or lower than it.
So far Altmetric has tracked 481 research outputs from this source. They typically receive more attention than average, with a mean Attention Score of 8.0. This one is in the 17th percentile – i.e., 17% of its peers scored the same or lower than it.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 263,642 tracked outputs that were published within six weeks on either side of this one in any source. This one is in the 34th percentile – i.e., 34% of its contemporaries scored the same or lower than it.
We're also able to compare this research output to 19 others from the same source and published within six weeks on either side of this one. This one is in the 10th percentile – i.e., 10% of its contemporaries scored the same or lower than it.