Voted Drug Discovery and Development Product for 2019: NanoBRET™ TE Kinase Assays

Posted on by Kari Kenefick
Choice Drug Discovery and Development Product 2019 award
Michael Curtin, Promega, accepting the Reviewers’ Choice for Drug Discovery and Development Product of the Year award from SelectScience.

As announced at SLAS in Washington, D.C. recently, we are excited to have NanoBRET™ Target Engagement (TE) Intracellular Kinase Assays awarded the SelectScience Reviewers’ Choice for Drug Discovery and Development Product of the Year 2019!

The NanoBRET™ Target Engagement (TE) Kinase Assay, first available in the fall of 2017, has been getting great reviews on the SelectScience site for more than a year now.

We are grateful to have been selected for this award and for reviewers’ votes and comments! Some of the comments include:

 

Review of NanoBRET TE Kinase Assay

Reviewer's comment on NanoBRET TE Kinase Assay

 

 

 

 

 

NanoBRET TE Kinase reviewers comment

 

 

 

Here are some details on what this assay can do as a tool in your drug discovery and development research.

What is the NanoBRET™ TE Kinase Assay ?
NanoBRET™ TE Intracellular Kinase Assays are the first biophysical method to enable the quantitative determination of compound potency and binding affinity for specific kinase targets inside live cells. The assays provide a direct measure of compound binding or occupancy to a target kinase under physiological conditions.

Ready-to-use target-specific cellular assays included in the NanoBRET™ TE Kinase family have been developed for >200 full-length kinases, including integral membrane receptor tyrosine kinases, as well as kinases that rely on regulatory proteins like the cyclin-dependent kinases.

All assays use a simple protocol that is conducted in multiwell plates and is scalable from 96-well to 384-well formats. The data quality from these NanoBRET™ TE Kinase Assays is excellent, with low error rates and high reproducibility.

Cellular potency of multiple kinase inhibitors types (I, II and allosteric compounds) have been measured using NanoBRET™ TE Kinase assays. Additionally, the NanoBRET™ TE Kinase assays are versatile and can be configured for kinetic or residence time analysis that enables the investigation of the duration of binding between a compound and a specific kinase in live cells.

Learn more about the NanoBRET™ TE Intracellular Kinase Assays at this product page, where you’ll find data, the protocol and a paper on how these assays have been applied to kinase profiling in live cells.

We hope you’ll find NanoBRET™ TE Kinase Assays a useful, exciting tool in your drug discovery and development workflow.

The Secret Fluorescent Life of Flying Squirrels

Posted on by Mariel Mohns
flying squirrel specimen
A flying squirrel museum specimen under normal light versus ultraviolet light. Photo courtesy of AM Kohler, et al.

In May 2017, a surprising discovery was made in the woods of Bayfield County, Wisconsin, just about a 5-hour drive north of Promega headquarters. Jonathan Martin, Associate Professor of Forestry at Northland College, was exploring the forest with an ultraviolet (UV) light in search of fluorescent lichen or plant life. What he found instead was a bright pink glow coming from a most unexpected source—a flying squirrel.

Continue reading “The Secret Fluorescent Life of Flying Squirrels”

How Gut Microbes Affect Our Brain

Posted on by Johanna Lee

coliform bacteriaThink about the last time you gave a presentation. The feeling of having “butterflies in your stomach”. Or when you meet someone for the first time, that “gut feeling” of whether you two will get along. In our day-to-day lives, we often associate what happens in the gut with what goes on in our brain. In fact, scientific evidence suggests that our gut and our brain frequently communicate—through gut microbes. Apparently, the existence of trillions of bacteria and eukaryotes in our gut is not only crucial for our physical health, they may also be important for our mental health.

Continue reading “How Gut Microbes Affect Our Brain”

It’s Almost iGEM Season—Help Is On The Way!

Posted on by Darcia Schweitzer

The 2019 iGEM Competition is on the horizon and team registration opens this month. We’re excited to partner with the iGEM Foundation again this year and offer our support to the young scientists who participate. If you’re starting an iGEM project, there are going to be things you need along the way. We are pleased to share a number of different ways we can help your iGEM team from now until the Giant Jamboree.

Grant Sponsorship

Tell us about your iGEM project and your team could win a 2019 Promega iGEM Grant Sponsorship. Ten winning teams will each receive $2000 in free Promega products to use for their iGEM projects. Tell us about your project—What problem are you addressing? What is your proposed solution? What challenges does your team face? Last year’s winning teams selected from a wide range of reagents and supplies, including master mix, restriction enzymes, ligase, DNA purification kits, expression systems, DNA ladders and markers, buffers and agarose. Click here to apply! Continue reading “It’s Almost iGEM Season—Help Is On The Way!”

Executing a NanoBRET™ Experiment: From Start to Data

Posted on by Promega

This is a guest post from Katarzyna Dubiel, marketing intern in Cellular Analysis and Proteomics.

“The objective of my experiment was to test the NanoBRET™ assay as if I was a customer, independent of the research and development team which develops the assay.”

Designing and implementing a new assay can be a challenging process with many unexpected troubleshooting steps. We wanted to know what major snags a scientist new to the NanoBRET™ Assay would encounter. To determine this, we reached out to Laurence Delauriere, a senior applications scientist at Promega-France, who had never previously performed a NanoBRET™ assay. Laurence went step-by-step through the experimental process looking at the CRAF-BRAF interaction in multiple cell lines. In an interview, Laurence provided us with some tips and insights from her work implementing the new NanoBRET™ assay.

In a few words, can you explain NanoBRET?
“NanoBRET is used to monitor protein: protein interactions in live cells. It is a bioluminescence resonance energy transfer (BRET) based assay that uses NanoLuc® luciferase as the BRET energy donor and HaloTag® protein labeled with the HaloTag® NanoBRET™ 618 fluorescent ligand as the energy acceptor to measure the interaction of two binding partners.” Continue reading “Executing a NanoBRET™ Experiment: From Start to Data”

Optimizing Pressure Cycling Sample Preparation for Bottom-Up Proteomics

Posted on by Gary Kobs

Large-scale analyses of the proteome have revealed proteomic changes in response to disease, and these changes hold great promise for diagnostics and treatment of complex disease if proteomic analysis can be brought into the clinical laboratory. Successful and reliable large-scale proteomics requires sample preparation workflows that are reproducible, reliable and show little variability. To bring proteomics into the clinical laboratory, standardized procedures and workflows for sample prep and analysis are required to generate valid, actionable results on a time scale useful for the clinic.

The two most common sample types analyzed for clinical proteomics are body fluids and tissue biopsies. To process these kinds of samples, there are two initial steps: tissue solubilization, followed by proteolytic digestion. Solubilization of solid tissues is the most labor-intensive and produces the most variable results.

The introduction of pressure cycling technology (PCT) using Barocycler instrumentation has greatly improved both tissue solubilization and digestion consistency. The PCT-based sample preparation protocols generally utilize urea as a lysis buffer for protein denaturing and solubilization. Urea has several drawbacks including inhibiting trypsin activity and introducing  unwanted modifications like carbamylation.

Lucas and colleagues analyzed whether replacing urea with SDC would produce similar tissue digestion profiles and improve the PCT method.

SDC allowed the use of higher temperatures compared to urea, and hence the first step (lysis, reduction, and alkylation) was performed at 56 °C. The second digestion step in the Barocycler was optimized, and the third step was eliminated. To further reduce digestion time, they capitalized on Rapid Trypsin/Lys-C.  Rapid Trypsin/Lys-C maintains robust activity at 70 °C, and allowed Barocycler digestion to be performed in a single step, completing digestion in 30 cycles (approximately 30 min) rather than 105 minutes, streamlining the protocol.

The data presented an improved conventional tissue PCT approach in a Barocycler by replacing urea and proteolytic enzymes with SDC, N-propanol, and modified commercially available enzymes that have higher optimum temperatures.

Paper Referenced

Lucas, N. et al. (2019) Accelerated Barocycler Lysis and Extraction Sample Preparation for Clinical Proteomics by Mass Spectrometry. J of Proteome Res 18, 399–405.

Expanding the Plague Family Tree: Yersinia pestis in the Neolithic

Posted on by Sara Klink
Yersinia pestis. See page for author [Public domain], via Wikimedia Commons

In recent years, scientists have been able to refine their molecular tools to resurrect ancient DNA from human graves and determine that yes, Yersinia pestis was the causative agent for the Black Death in the 14th century and the Plague of Justinian in the 6th century.  As more and more human graves have been uncovered, their DNA has revealed many secrets that scientists even ten years ago were unable to discover. With the ability to sequence entire genomes of bacteria that died with their hosts hundreds and even thousands of years ago, researchers are exploring the rise and possible spread of Y. pestis. Each new member sequence adds to the Y. pestis family tree, pinpointing the origin of this bacteria as it diverged from its ancestor Y. pseudotuberculosis. Peering into the past, scientists have been able to track down a strain of Y. pestis from individuals in a Swedish passage grave that is basal to known strains and that the authors of a Cell article suggest has interesting implications.

This pathogenic journey into history started by analyzing ancient DNA data sets from the teeth of individuals present in a communal passage grave in Gökhem parish, located in western Sweden, for any disease-causing microbial sequences that might be present. Y. pestis was flagged in one 20-year-old female dated 4,867–5,040 years ago. The bacterial sequences from this individual, named Gok2, were more closely aligned with Y. pestis than the Y. pseudotuberculosis reference genome. Continue reading “Expanding the Plague Family Tree: Yersinia pestis in the Neolithic”

Extra extra: Read All About Tautonyms

Posted on by Julia Nepper

If you’re active on #sciencetwitter, you may have seen a thread recently about tautonyms. “Tautonym” is a cool word for scientific names where the genus and species are the same word, For example, Vulpes vulpes is the scientific name for the red fox.

I have taken great delight in sharing these tautonyms with friends, colleagues, and random strangers on the bus. However, the problem that I keep having is that people want more details about something than the name. If you’ve had that problem, too, then this blog is for you. Continue reading “Extra extra: Read All About Tautonyms”

Goodbye to the Most Famous Bird in Maine

Posted on by Jordan Villanueva

When Wisconsin plunged into a deep freeze during last week’s polar vortex, I built a roaring fire in my fireplace and settled into my armchair with a thick blanket and a video game controller. Except for the twenty minutes I spent driving to and from the office, I stayed warm and toasty.

Birds, however, don’t have it quite as easy. To survive freezing temperatures, non-migratory birds have developed many interesting adaptations. Many species grow extra down layers and huddle together for wind protection. Others, like the black-capped chickadee, use a process called regulated hypothermia to drop their resting body temperature by as much as 22°F to conserve energy. I’m particularly fascinated by the process of regional hypothermia—many species of ducks and gulls use a countercurrent heat exchange system to keep vital organs warm while letting temperatures fall in extremities.

Birds that aren’t accustomed to cold weather don’t have these adaptations, though. When a bird—or any animal—ends up far outside of its natural habitat, the consequences can be deadly.

Continue reading “Goodbye to the Most Famous Bird in Maine”

Oh, the Places You’ll Go, Thanks to Science!

Posted on by Promega

Today’s guest blog is written by Aparna Shah, a Post-Doc at Johns Hopkins University. Aparna visited the Promega campus in Madison, Wisconsin on January 18, 2019 and offered to share her experiences.

Aparna poses with her artwork at the Promega Employee Art Showcase

I don’t recall ever having won a contest before, let alone the grand prize! In fact, I did a double take when I first read the email informing me that my SciArt submission had been selected as a winning entry for the Promega Art Contest for Creative Scientists. What did I win, you ask? A free trip to Madison, WI to meet with the team behind the contest and explore Promega’s headquarters!

I first heard about the contest on the HelloPhD podcast and considered participating primarily to support the SciArt movement. A couple of days later, I came across a perfectly-timed tweet about the contest that nudged me out of procrastination mode and reminded me to follow through with it. I’m going to take a second here to pitch both HelloPhD and Twitter to you. Regardless of whether you’re an undergraduate student interning in a science lab or a senior postdoc, the HelloPhD podcast is incredible at calming you down while you’re on the roller coaster ride called academia. As for Twitter, I can think of several pros for using it. But in the context of this post, it is one of the best resources for discovering opportunities that match your interests.

Continue reading “Oh, the Places You’ll Go, Thanks to Science!”