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Flow cytometric measurement of the cellular propagation of TDP-43 aggregation

Prion. 2017-05; 
ZeineddineRafaa, WhitenDaniel R, FarrawellNatalie E, McAlaryLuke, HanspalMaya A, KumitaJanet R, WilsonMark R, YerburyJust
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Mutagenesis Services … TDP-43-tomato red (TDP-43-tdTo- mato) constructs were created by replacing the tGFP sequences with tdTomato (Genscript, USA); site-directed mutagenesis was also per- formed by Genscript (USA) to create the G294A mutant (TDP-43G294A) … Get A Quote

摘要

Amyotrophic lateral sclerosis is a devastating neuromuscular degenerative disease characterized by a focal onset of motor neuron loss, followed by contiguous outward spreading of pathology including TAR DNA-binding protein of 43 kDa (TDP-43) aggregates. Previous work suggests that TDP-43 can move between cells. Here we used a novel flow cytometry technique (FloIT) to analyze TDP-43 inclusions and propagation. When cells were transfected to express either mutant G294A TDP-43 fused to GFP or wild type TDP-43fused to tomato red and then co-cultured, flow cytometry detected intact cells containing both fusion proteins and using FloIT detected an increase in the numbers of inclusions in lysates from cells expre... More

关键词

ALS,FloIT,Flow cytometry,Prion,Propagation,Protein Aggregation,TD