Single Cell Western Testimonials

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<div class="highlighted"> <aside class="container section clearfix" role="complementary"> <div> </div> </aside> </div> <div id="main" class="container-fluid container-wide "> <div class="row"> <div class="col-md-12"> <div> <div id="block-bio-techne-global-breadcrumbs" class="system_breadcrumb_block"> <nav role="navigation" aria-labelledby="system-breadcrumb"> <h2 id="system-breadcrumb" class="visually-hidden">Breadcrumb</h2> <ol> <li> <a href="/">Home</a> </li> <li> <a href="/resources">Resources</a> </li> <li> <a href="/resources/testimonials">Testimonials</a> </li> <li> Milo Goes Keto: Conquering Analysis of Scarce Intestinal Stem Cell Populations </li> </ol> </nav> </div> </div> </div> </div> <div class="row"> <div class="col-md-12"> <main> <section class="section"> <div> <div id="block-bio-techne-global-content" class="system_main_block"> <article> <h1><span>Milo Goes Keto: Conquering Analysis of Scarce Intestinal Stem Cell Populations</span> </h1> <div class="testimonial_published_dt"> Posted August 10, 2021 </div> <div> <div class="container card testimonial mb-5 p-3"> <div class="row"> <aside class="col-lg-3 text-center m___testimonial-aside"> <img src="https://resources.bio-techne.com/bio-techne-assets/images/testimonials/milo-single-cell-western-chia-wei-cheng.png" alt="Chia-Wei Cheng, Ph.D." loading="lazy" > <p class="text-center mt-2 font-weight-bold testimonial__profile"> Chia-Wei Cheng, Ph.D.<br/> Postdoctoral Fellow, Koch Institute for Integrative Cancer Research, MIT </p> </aside> <section class="col-lg-9 m___testimonial-section"> <div class="testimonial__quote"> <p class="font-italic">"With Milo, we observed a level of single-cell protein expression heterogeneity in our purified intestinal stem cell population that was not previously possible with conventional Western blotting."</p> <p>- Chia-Wei Cheng, Ph.D., Postdoctoral Fellow, Koch Institute for Integrative Cancer Research, MIT</p> </div> <div> <div> <div> <div class="paragraph paragraph--type--freeform mb-4 paragraph--view-mode--default"> <div><h3> </h3> <h3>IN THE CRYPTS OF DIETARY COMPOSITION</h3> <p>Obesity and related co-morbidities are a major global health problem. On the individual level, dietary composition is a key contributor to health and disease—influencing metabolic changes that either fuel tissue regeneration and slow aging or impair tissue homeostasis and promote obesity and cancer. To that end, adult intestinal stem cells (ISCs), located toward the base of the crypts of the adult mammalian intestine, both mediate the renewal of the intestinal epithelium and orchestrate the molecular response to dietary and energy cues. Still, the stem and progenitor cell biology, as it relates to diet perturbation, is not well characterized. The research laboratory of Ömer H. Yilmaz, M.D, Ph.D., at the Koch Institute for Integrative Cancer Research at MIT is focused on furthering our understanding of adult ISCs. Specifically, they study how ISCs and their microenvironment adapt to diverse diets in the context of tissue regeneration, aging, and cancer initiation/progression.</p> <p> </p> <h3>KETOGENIC DIETS AND HEALING FROM WITHIN</h3> <p>Chia-Wei Cheng, Ph.D., a Postdoctoral Fellow in the laboratory of Dr. Yilmaz, studies dietary control of adult ISC fate and tissue maintenance. She is also the lead author of a recent study in <em>Cell</em> showing that a ketogenic diet in mice gives ISCs a regenerative boost, compared with mice on a regular diet. The underlying mechanisms are attributed to a dietresponsive metabolite signaling axis that modulates the well-known Notch signaling pathway to enhance stem cell renewal and function.</p> <p>But Dr. Cheng’s work and recent accomplishments have not come without technical challenges. Adult stem cells in the intestinal epithelium are rare (~1–3%) and exist in a mixed population with neighboring Paneth cells and deep secretory cells of the colon. Cell sorting according to the leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5) marker is necessary to obtain a pure population for accurate conclusions. In identifying potential metabolic pathways augmented in ISCs, Dr. Cheng and colleagues could turn to RNA-seq data to look at differential gene expression in populations of flow-sorted Lgr5-GFPhr ISCs.</p> <p>However, her efforts became stymied by the lack of a highly sensitive, sample-preserving technical approach for confirmation of her RNA-seq results on a protein level. "Because of their scarcity, sorting ISCs for Western blotting has been very time consuming, and also, the number of total allowable tests per animal is very limited. Conventional Western blotting requires at least 50,000 cells to determine the expression levels of each protein," Dr. Cheng says about her day-to-day. But adds cheerfully, "We decided to implement Milo to validate the results of gene expression analyses for Lgr5+ ISCs."</p> <p> </p> <h3>MILO RELIABLY MEASURES PROTEIN EXPRESSION IN LOW ABUNDANCE SAMPLES</h3> <p>The <a href="/p/single-cell-western/milo_p100">Milo™</a> <a href="/instruments/single-cell-western">Single-Cell Western</a> system provides quantitative information in two output formats: by the measurement of protein target abundance (<em>i.e.</em>, signal) in every single cell and by the enumeration of target positive cells. With Milo, the number of cells captured scales with the number of cells loaded, so even lower abundance samples are possible. "Milo has enabled us to use less than 1,000 cells to answer a question that otherwise demanded a minimum of 50,000 cells to do so," attests Dr. Cheng. The answer to this question—whether or not a key mitochondrial enzyme involved in ketogenesis was enriched in Lgr5+ crypt ISCs—allowed the team to make functional conclusions about this stem cell population.</p> <p>Using Milo, Dr. Cheng was able to measure 3-Hydroxy-3-methylglutaryl-CoA synthase 2 (Hmgcs2) protein expression levels at single-cell resolution. More specifically, the team could compare the Hmgsc2/β-tubulin level per cell in total epithelial cell populations versus that found in flowsorted Lgr5high ISCs. Consistent with their omics data, Single-Cell Westerns on Milo also illustrated that Hmgcs2-expressing cells (Hmgcs2+) were highly enriched in flow-sorted Lgr5+ ISCs (77.97%) but less frequent in total intestinal crypt cells (21.17%)<sup>1</sup>.</p> <p>The MIT team performed Single-Cell Western blotting experiments on Milo using the Standard <a href="/search?keywords=scWest%20Kit&view=condensedCell%20Western">scWest kit</a> and Scout™ Software, which not only provides quantitative results but automates the data analysis workflow!</p></div> </div> </div> </div> </div> <div class="text-center p-2" style="background-color:#f2f2f2;"> <h3>Request a Quote</h3> <p> Get a personalized quote for your Single-Cell Western instrument </p> <p> <a href="/p/instruments/single-cell-western/request-quote" class="btn btn-primary">Request Quote</a> </p> </div> <div> <h4 class="mt-3">Selected Publications</h4> <div> <div id="accordion" class="paragraph mb-4 paragraph--type--references-container paragraph--view-mode--default"> <div class="card-header"> <a class="" data-toggle="collapse" data-target="#refPara-36826" role="button" aria-expanded="true" aria-controls="refPara-36826"> <h5 class="d-inline">References</h5> </a> </div> <div id="refPara-36826" class="paragraph mb-4 paragraph--type--references-container paragraph--view-mode--default collapse accordion show" aria-controls="refPara-36826"> <ol class="py-3"> <li class="py-2"> <p class="paragraph paragraph--type--individual-reference paragraph--view-mode--default image-and-text-default"> <strong>CW Cheng</strong> <em> et al. </em> (2019) <a href="https://www.ncbi.nlm.nih.gov/pubmed/31442404" title="View Abstract" target="_blank" rel="noreferrer"> Ketone body signaling mediates intestinal stem cell homeostasis and adaptation to diet</a> Cell <strong>178</strong>:1115-31. PMID: 31442404. </p> </li> </ol> </div> </div> </div> </div> </section> </div> </div> </div> </article> </div> </div> </section> </main> </div> </div> </div> <div id="brandsbar-footer" class="bg-bt-primary"> <div class="container-wide container-fluid text-white"> <div class="row py-5"> <div class="footer-logo col-xl-3 col-lg-auto d-flex"> <img class="brand-bar-footer-logo" src="/themes/custom/bio_techne_global/logo.svg" alt="Bio-techne logo"> </div> <div class="col-xl-9 col-lg-auto"> <p class="ml-xl-4 bt-px_groteskbold footer-tagline">Global Developer, Manufacturer, and Supplier of High-Quality Reagents, Analytical Instruments, and Precision Diagnostics.</p> </div> <div class="included-brands col"> <span class="font-weight-bold mr-3 bt-inter_regular">INCLUDES</span> <span class="mr-3 bt-inter_regular">R&D Systems™</span> <span class="mr-3 bt-inter_regular">Novus Biologicals™</span> <span class="mr-3 bt-inter_regular">Tocris Bioscience™</span> <span class="mr-3 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