Biotech Manufacturing: A Good Machinist is Critical for Your Laboratory Reagents

Travis Beyer, Machinist Technician, at the CNC milling machine in the Promega machine shop.

Travis Beyer, Machinist Technician, at the CNC milling machine in the Promega machine shop.

It can be easy to forget that Promega is a manufacturing business. The company’s cGMP Feynman Center on the Madison campus has been described as looking more like a retreat than a factory. But hidden within the well-designed walls of Feynman, as well as in other facilities on campus, technicians operate hundreds of machines that manufacture, dispense and package Promega reagents day in and day out. Keeping those high-tech machines running at peak performance is critical, requiring immense skill, precision and even artistry. That’s where Promega Machinist Technician Travis Beyer comes in.

“I get to make stuff,” says Travis who is not afraid to show his enthusiasm for his craft while describing the best part of his job. “There’s a product at the end of the day. Plus I get to support science, and make things that support people’s lives. That’s cool.”

 I get to make stuff. There’s a product at the end of the day. Plus I get to support science, and make things that support people’s lives. That’s cool.

The da Vinci Center, another artfully designed building on the Madison campus, houses the Promega machine shop where Travis does his work designing or improving on parts for newer manufacturing equipment or reverse engineering broken or worn parts no longer available for older equipment that still serves its purpose. He makes every machine part imaginable from drive shafts to sensor brackets to filling forks, and his work is critical to manufacturing businesses like Promega, where a downed piece of equipment can cause costly production delays.

An example of a machine part that Travis designs or reverse engineers and then builds to keep Promega manufacturing moving smoothly.

An example of a machine part that Travis designs or reverse engineers and then builds to keep Promega manufacturing moving smoothly.

As he explains, not many manufacturing companies the size of Promega have a fully capable machine shop. They usually send out their work, meaning longer lead times and more expense. But, as its distinctive architecture suggests, Promega is not like many other companies. Continue reading

Making BRET the Bright Choice for In vivo Imaging: Use of NanoLuc® Luciferase with Fluorescent Protein Acceptors

13305818-cr-da-nanoluc-application_ligundLive animal in vivo imaging is a common and useful tool for research, but current tools could be better. Two recent papers discuss adaptations of BRET technology combining the brightness of fluorescence with the low background of a bioluminescence reaction to create enhanced in vivo imaging capabilities.

The key is to image photons at wavelengths above 600nm, as lower wavelengths are absorbed by heme-containing proteins (Chu, J., et al., 2016 ). Fluorescent protein use in vivo is limited because the proteins must be excited by an external light source, which generates autofluorescence and has limited penetration due to absorption by tissues. Bioluminescence imaging continues to be a solution, especially firefly luciferase (612nm emission at 37°C), but its use typically requires long image acquisition times. Other luciferases, like NanoLuc, Renilla, and Gaussia, etc. either do not produce enough light or the wavelengths are readily absorbed by tissues, limiting their use to near- surface imaging.

The two papers discussed here illustrate how researchers have combined NanoLuc® luciferase with a fluorescent protein to harness bioluminescent resonance energy transfer (BRET) for brighter in vivo imaging reporters. Continue reading

Preventing Viral Infection by Blocking Cellular Receptors with a Tethered Antibody

Cross section of mature HIV. Copyright David S. Goodsell, The Scripps Research Institute.

Cross section of mature HIV. Copyright David S. Goodsell, The Scripps Research Institute.

Finding a way to neutralize or block infection by HIV has long been pursued by viral researchers. Various treatments have been developed, driven by the need to find effective drugs to manage HIV in infected individuals. The ultimate goal is to develop a vaccine to prevent HIV from even taking hold in the body. With all of our knowledge about the cellular receptors HIV needs to enter the cell, there has to be a method to prevent a viral particle from binding and being internalized. Many researchers are pursuing neutralizing antibodies to the virus as one method. What about antibodies that target the cellular receptor recognized by the virus? In a recently published article in Proceedings of the National Academy of Sciences, antibodies to cellular receptors for rhinovirus and HIV were tethered to the plasma membrane and tested for the ability to prevent infection. Continue reading

Don’t Let These Three Common Issues Hurt Your Luminescent Assay Results

4621CAThere is a lot riding on your luminescent assay results. Each plate represents precious time, effort and resources. Did you know that there are three things about your detection instrument that can impact how much useful information you get from each plate?  Instruments with poor sensitivity may cause you to miss low-level samples that could be the “hit” you are looking for.  Instruments with a narrow detection range limit the accuracy or reproducibility you needed to repeat your work.  Finally, instruments that let the signal from bright wells spill into adjacent wells allow crosstalk to occur and skew experimental results, costing you time and leading to failed or repeated experiments. Continue reading

Bones: Improved Technology is Bringing Loved Ones Home

21537407 - archeology, human bonesToday’s Promega Connections blog is written by guest blogger Rachel H. Oefelein, QA Manager/Senior DNA Analyst at DNA Labs International. 

Shakespeare said, “The evil that men do lives after them; the good is oft interred with their bones.”  This is continually true in the case of unidentified remains throughout the United States.  The action of a person going missing or the events leading to an individual’s demise are frequently the memory that haunts a town or the media for years to come. However, for each such case, somewhere lies a set of skeletal remains not yet found, or just as tragic, recovered but still unidentified.  The National Missing and Unidentified Persons System (NamUs) estimates approximately 40,000 sets of unidentified skeletal remains linger in morgues around the country or that have been cremated and buried as Jane and John Does.

Many crime labs do not have protocols in place for the extraction of DNA from skeletal remains or have outdated protocols for bone extraction that are not sensitive enough for poor quality bones. Bones are often recovered from harsh environments and have been exposed to extreme heat, time, acidic soil, swamp, chemicals treatment, etc. These harsh environmental conditions degrade the DNA present in the remains which further complicates the already difficult procedure of releasing the DNA in cells buried deep within the bone matrix. Another challenge is that cases often involve recovery of skeletal remains in areas with animal activity, water recoveries and scenes involving explosions or fires; these case types may require re-association of dozens if not hundreds of bones and bone fragments. Continue reading

Making a Case for Basic Research Funding

The value of public funding for “basic” versus “applied” research has long been questioned. To address this debate, the authors of a recent report in Science performed a large-scale evaluation of the value of public investment in biomedical research. After analyzing the relationship between the U.S. National Institutes of Health (NIH) grants and private patents, they found that distinguishing research as basic or applied is not useful in determining the productivity of grant funding.

Genetic research at the laboratoryThe $30 billion annual budget of the NIH makes it the largest source of life science funding in the world and provides a third of all US biomedical research and development. Although there has long been a strong argument for public investment in scientific research, attacks on the tangible benefits of this research persist. In particular, some opponents argue that “basic” research is too far removed from practical applications to be worthy of investment.

To quantify the effects of NIH funding for basic versus applied research, the authors looked at data from 365,380 grants awarded between 1980–2007 and compared their direct and indirect influence on patent filed. In particular, they decided to use patent-article citations as a measure of the influence of publicly funded science on commercial developments.

The researchers determined two ways in which research funded by the NIH could impact patenting; patents could be filed by the NIH-funded scientists or by private entities that cited research funded by NIH grants. This study found that roughly 10% of NIH grants were directly responsible for a patent while nearly a third of NIH grants had an indirect influence on patents. This indirect influence was attributed to articles associated with grant research that were later cited by a patent.

Delving deeper into the data, the authors found a similar pattern when looking at drugs brought to market that were associated with NIH grants; less than 1% of grants were directly linked to a patent associated with a drug, while 5% resulted in a publication cited by a patent for a drug. Despite public policies like the Bayh-Dole Act, that encourage academic researchers to file their own patents, the traditional route of applying public research to private patents continues to predominate.

For those that question the value of basic research and aim to steer public policy toward supporting applied research, this report makes a strong case against this way of thinking. The findings also suggest that using direct generation of patents as a metric for the return on investment of publicly funded biomedical research is not very useful since most of the effects of NIH research appear to be indirect.

In fact, the authors posit that basic research is just as productive as applied research in terms of patenting since the amount of grant research cited by private patents is much greater than the number of grants directly associated with patents. Perhaps it is time policy makers consider studies like this and forgo disseminating grant funds based on whether research is basic or applied.

The Randomness of Cancer

A major scientific study grabbed headlines recently, and the implications of its findings may affect many of us, if not all of us. In a paper published in Science by Cristian Tomasetti, Lu Li and Bert Vogelstein of Johns Hopkins University, the authors report that nearly two-thirds of known cancer causing mutations can be attributed to random mistakes that occur during DNA replication. In other words, the vast majority of these mutations occur in a spontaneous, uncontrollable way— it may not matter how you live your life, or what measures you take to decrease your chance of developing cancer. As the authors and the press put it, it really just comes down to luck.

gene-mutationDisturbing? For many, yes. It’s not easy to accept that one’s luck in activities such as winning the lottery may also apply to whether or not you will be touched by cancer. That is partly why this study is gaining so much attention.

As the authors explain in their publication, until now most cancer-causing mutations had been attributed to two major sources: inherited and environmental factors. But they found that a third kind of mutation, replicative (R) mutations that arise from unavoidable errors associated with DNA replication, account for 66 percent of mutations that drive cancer. Continue reading

Researchers Gather at Promega Madison Campus for Annual Stem Cell Symposium

stemcell header

Since the derivation of human-derived embryonic stem cells (ES cells) in the 1990’s, the world of stem cell biology and engineering has proceeded at an amazing pace. The isolation pluripotent cells (iPS) cells that have most of the properties of embryonic stem cells from somatic tissues has been possible for nearly a decade. Engineered human cells, tissues, and organ-like structures are becoming a reality and may soon play a part in treating diseases. ES and iPS cells are teaching us much about how cells become specialized during normal development and the pathologies that result when those specialization decisions go wrong.

At the 12th Annual Wisconsin Stem Cell Symposium held at the BioPharmaceutical Technology Center institute, leading researchers from around the world will be gathered to discuss the latest progress, roadblocks and issues around Engineering Cells and Tissues for Discovery and Therapy.

The Symposium is co-coordinated by the Stem Cell & Regenerative Medicine Center at the University of Wisconsin-Madison and the BioPharmaceutical Technology Center Institute and is open to the public. Registration is $100.00 ($50.00 for students and post-doctoral researchers). The Symposium will be held at Promega Corporation’s BioPharmaceutical Technology Center, 5445 E. Cheryl Parkway, Fitchburg, WI.

Topics to be discussed include: Continue reading

Meet Katie Herbrand, Genetic Identity Supervisor on the Spectrum Team

29160613_lPromega will introduce the Spectrum CE System for forensic and paternity analysis. Building this system requires the efforts of many people from many disciplines–from our customers who have told us their needs to the engineers and scientists building the instrument and ensuring its performance. Periodically we will introduce our Promega Connections readers to a team member so that you can have a sneak peak and behind-the-scenes look at Spectrum CE System  and the people who are creating it (of course if you truly want to be the first to know, sign up at www.promega.com/spectrum to receive regular, exclusive updates about Spectrum CE).

Today we introduce Katie Herbrand, Genetic Identity Supervisor in Production. Continue reading

Making the Switch from FRET to BRET: Applications of NanoLuc® Luciferase with Fluorescent Protein Acceptors for Sensing Cellular Events

A Bioluminescent Alternative

Fluorescence resonance energy transfer (FRET) probes or sensors are commonly used to measure cellular events. The probes typically have a matched pair of fluorescent proteins joined by a ligand-binding or responsive protein domain. Changes in the responsive domain are reflected in conformational changes that either bring the two fluorescent proteins together or drive them apart. The sensors are measured by hitting the most blue-shifted fluorescent protein with its excitation wavelength (donor). The resulting emission is transferred to the most red-shifted fluorescent protein in the pair, and the result is ultimately emission from the red-shifted protein (acceptor).

As pointed out by Aper, S.J.A. et al. below, FRET sensors face challenges of photobleaching, autofluorescence, and, in the case of exciting cyan-excitable donors, phototoxicity. Another challenge to using FRET sensors comes when employing optogenetic regulators to initiate the event you wish to monitor. Optogenetic regulators respond to specific wavelengths and initiate signaling. The challenge comes when the FRET donor excitation overlaps with the optogenetic initiation wavelengths. Researchers have sought to alleviate many of these challenges by exchanging the fluorescent donor for a bioluminescent donor, making bioluminescence resonance energy transfer (BRET) probes. In the three papers described below, the authors chose NanoLuc® Luciferase as the BRET donor due to its extremely bright signal. Continue reading