Miranda Sets a New Pace for Alzheimer's Research with Wes

FIGHTING TO KEEP OUR WITS

Miranda Orr is a Postdoctoral Fellow at the Barshop Institute for Longevity and Aging Studies, University of Texas Health Science Center in San Antonio, where researchers study the basic biology of aging. They use animal models from invertebrates to non-human primates to study the cellular and molecular driving forces responsible for "aging". They also test the utility of pharmacological, dietary, and physical interventions to extend the healthy years of life. Miranda's research interest centers on understanding the pathophysiology of Alzheimer's disease and the mechanisms governing the divergence of normal brain aging toward a progressive, irreversible pathogenic state. Although she's always been innately curious about brain function and aging, her research is extremely personal too. She pursued a Ph.D. in neuroscience after watching her vibrant and quickwitted grandmother slowly lose her memories after being diagnosed with Alzheimer's disease. Her goal? Make a big difference in the lives of the aging by positively influencing Alzheimer's disease research.

ALL HANDS-ON, ALL THE TIME

Miranda studies how age-associated changes in normal physiology alter the expression and function of the microtubule associated protein tau. Tau is highly expressed in neurons and its misprocessing occurs in many neurodegenerative disorders, including Alzheimer's disease. Tau phosphorylation negatively regulates its microtubule binding function, and sustained high levels of phosphorylation can lead to a toxic gain of function, neuronal dysfunction and degeneration. Traditional Western blot proved to be more than a hefty task given she was comparing multiple brain regions across different experimental changes like drug intervention and dietary change, and across lifespan, and her protein of interest (tau) can gain extensive posttranslational modifications at dozens of unique sites. She had all the latest equipment for high throughput Western blotting like pre-cast gels, 7-minute transfer methods, IR-dye conjugated secondary antibodies for multiplexing and a high-end Imaging system. But even with all that she felt like a processing slave with all the hands-on time. And after all the data were collected, she still had to tackle analysis, which was just as time-consuming. On top of that, low abundant proteins or small brain sub-regions were limiting how many data points she could get from any particular sample.

CLEARING THE SMEAR

Wishing for a "simpler" Western, Miranda tried Wes. He let her run 24 independent samples and get fully analyzed data in about 4 hours. She also multiplexed up to 5 antibodies per capillary to get 120 independent data points in those same 4 hours. She found Wes reproducible, reliable, fast, and used 95% less tissue and antibody. He also freed up her time for other experiments. So she started testing more proteins along with additional samples, experimental conditions and brain regions, and did it all in the same time it would take with traditional Western blots or less! The lab's newly upped capacity also helped Miranda discover a novel high molecular weight isoform of tau protein that is expressed in the brains of the naked mole-rat (NMR), the longest living rodent known. She was perplexed by a higher molecular weight protein smear she kept seeing in her blots. But when she ran the same samples on Wes, she saw discrete high molecular weight bands much larger in size than any other brain tau isoform she'd studied before. The results showed that tau undergoes a progressive shift in molecular weight during the first year of NMR brain development (Figure 1, M.E. Orr et al, Neurobiology of Aging 36, 2015).

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