Author: Kari Kenefick

Illuminating the Brain with a New Bioluminescence Imaging Substrate

Posted on by Kari Kenefick

Bioluminescence imaging is a powerful tool for non-invasive studies of the effect of treatments on cells and tissues. The luminescent signal is strong, and can be used in vivo, enabling repeated observations over time, allowing longitudinal study of cellular changes for hours or days. Bioluminescence imaging can be used in live animals over varying periods of time, without interfering with normal cellular processes.

Fluorescence imaging is also used in cellular studies. Although it can provide a stronger signal than luminescence, fluorescence requires light for excitation, and thus its in vivo use is limited at a tissue or cell depth greater than 1mm.

NanoLuc® Luciferase. Small, bright and now useful in brain bioluminescence imaging.

In addition, autofluorescence can be an issue with fluorescence imaging, as cellular components and surrounding proteins and cells can fluoresce when exposed to light. Autofluorescence can result in high background signals, making it difficult to distinguish true fluorescence from background.

Continue reading “Illuminating the Brain with a New Bioluminescence Imaging Substrate”

Made Just for You: Promega Custom Reagents

Posted on by Kari Kenefick

At promega.com you’ll find reagents designed for use in your life science research, whether you need to isolate DNA or RNA, determine cell viability or signaling, gain metabolic assay insights, run a reporter bioassay or isolate nucleic acid from wastewater.

Did you know that we also create unique custom reagents? Whether you’re looking for an extra-large size of a compound, a unique type of packaging or package labeling, or a reagent or assay target that’s unique to your project, Promega custom reagents can help.

What Types of Custom Reagents Are Available?
We currently supply custom-made reagents in the areas of amplification, bioluminescence, nucleic acid purification, protein analysis and protein purification. See this Promega Custom Products and Technologies web page for details.
Need a unique master mix for your amplification reaction? This short video provides examples of how we can customize an amplification master mix.

Continue reading “Made Just for You: Promega Custom Reagents”

Real-Time Analysis for Cell Viability, Cytotoxicity and Apoptosis: What Would You Do with More Data from One Sample?

Posted on by Kari Kenefick

Originally posted May 25, 2017. Updated 2022

You are studying the effects of a compound(s) on your cells. You want to know how the compound affects cell health over a period of hours, or even days. Real-time assays allow you to monitor cell viability, cytotoxicity and apoptosis continuously, to detect changes over time.

Why use a real-time assay?
A real-time assay enables you to repeatedly measure specific events or conditions over time from the same sample or plate well. Repeated measurement is possible because the cells are not harmed by real-time assay reagents. Real-time assays allow you to collect data without lysing the cells.

Advantages of  Real-Time Measurement
Real-time assays allow you to:

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Evidence of Inflammasome Activation in Severe COVID-19

Posted on by Kari Kenefick

The pandemic caused by SARS-CoV-2 has brought the world to its knees. There have been many deaths, many persons with lingering disease (long COVID) and the inability to vaccinate everyone quickly, for starters. SARS-CoV-2 has not only been a tricky adversary in terms of treatment options to save lives, it’s also been a wily opponent to researchers studying the virus.

Contributing to the existing studies, with their review of the role of inflammasomes in COVID-19, Vora et al. recently published “Inflammasome activation at the crux of severe COVID-19” in Nature Reviews Immunology. In this paper they detail evidence of inflammasome activation and its role in SARS-CoV-2 infections.

Contributions of Those Lost in the SARS-CoV-2 Pandemic
I’d like to take a moment to note the uniquely awful nature of the virus at the center of this blog and the paper it reviews. Many of the papers we blog about describe research involving cell lines, mice or another animal model. The closest most reports get to human research subjects is the use of human cells lines. In the Vora et al. report, serum and tissue samples are from actual human patients, some that survived and many that did not survive COVID-19. It’s not lost on us, Dear Reader, the contributions of those that suffered and died due to SARS-CoV-2 infection. Many persons with severe or fatal COVID-19 have made a significant contribution to our understanding of this virus and its treatment options. We owe them, as well as the researchers that have studied SARS-CoV-2, our sincerest gratitude.

Why the Interest in Inflammasomes?
For detailed information on inflammasomes you can read Ken’s blog, here. You will find background information there and on our inflammasome web page.

In their paper, Vora et al. provide evidence of inflammasome activation, both direct and indirect, in COVID-19. The authors note:

“Key to inflammation and innate immunity, inflammasomes are large, micrometrescale multiprotein cytosolic complexes that assemble in response to pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs) and trigger proinflammatory cytokine release as well as pyroptosis, a proinflammatory lytic cell death.”

Continue reading “Evidence of Inflammasome Activation in Severe COVID-19”

Psilocybin as Antidepressant: Quick Acting, Lasting Benefits

Posted on by Kari Kenefick

Depression is not simply a mood disorder, a feeling of sadness, or being ill at ease. Depression can completely shut a person down, manifesting as an inability to make decisions, to take action, to think. Even sleep is affected by depression.

Researchers and clinicians who treat depression are learning that the physical manifestations can be mirrored by internal, cellular changes. Some people with depression have decreases in their gray matter volume, particularly in areas like the hippocampus (important to memory, learning, and emotions) and prefrontal cortex (where higher-level thought and planning abilities are based).

Additionally, imaging has shown a decrease in the number of synapses—the structures through which electrical or chemical signals are passed between neurons and other cells—in persons with chronic depression. Without the signals that synapses transmit, brain function is disrupted.

And without intervention in depression, synapse decrease can continue.

While there are drugs and behavioral therapies to treat depression, these therapies can be slow to act and sometimes ineffective. In addition, once synaptic loss has occurred, these therapies are less effective.

In their August 2021 paper, “Psilocybin induces rapid and persistent growth of dendritic spines in frontal cortex in vivo” (1), Shao et al. state,

“It has long been recognized that these compounds (serotonergic psychedelics like psilocybin) may have therapeutic potential for neuropsychiatric disorders, including depression, obsessive-compulsive disorder and addiction”.

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Antibody Response Differ in Adults, Kids and Potential Cross-Reactive Coronavirus Antibodies

Posted on by Kari Kenefick
B cell and B cell receptor cartoon. The B cell receptors are important for antibody response.
Drawing of a B cell and the B cell receptor. The receptor shows the characteristic Y shape of an immunoglobulin molecule.

B cells are the immune cells that produce antibodies (immunoglobulins or Ig) to detect intruding pathogens. B cells produce a variety of classes of antibodies. Generally during an immune response to a pathogen, whether viral or bacterial, B cells produce immunoglobulins (Ig) IgM and IgD, and later in the response, IgG and IgA, that are specific to the intruding organism. These Igs capture and aid in neutralizing the pathogen.

Ig classes can be studied by sequencing the B cell receptor (BCR), which binds antigen specifically. BCRs are formed via irreversible gene segment rearrangements of variable, diversity and joining (VDJ) genes. Ig classes can be diversified through somatic hypermutation and class-switch recombination of these gene segments (1).

B cell receptors with high sequence similarity can be found in individuals exposed to the same antigen, demonstrating that antigen exposure can result in similar B cell clones and memory B cells between individuals, both adults and children (1).

However, B cell immune responses can differ between adults and children. For example, children use more B cell clones that form neutralizing antibodies to HIV-1. And children infected with SARS-CoV-2 generally have milder illness than infected adults. SARS-CoV-2-infected children have lower antibody titers to the virus and more IgG-specific response to SARS-CoV-2 spike protein than to the nucleocapsid protein (1). These differences can contribute to faster SARS-CoV-2 clearance and lower viral loads in children versus adults.

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NAD: A Renaissance Molecule and its Role in Cell Health

Posted on by Kari Kenefick

Promega NAD/NADH-Glo system and how to prepare samples for identification of NAD or NADH.
Promega NAD/NADH-Glo system and how to prepare samples for identification of NAD or NADH.

NAD is a pyridine nucleotide. It provides the oxidation and reduction power for generation of ATP by mitochondria. For many years it was believed that the primary function of NAD/NADH in cells was to harness and transfer energy  from glucose, fatty and amino acids through pathways like glycolysis, beta-oxidation and the citric acid cycle.

Today, however, NAD is recognized as an important cell signaling molecule and substrate. The many regulatory pathways now known to use NAD+ in signaling include multiple aspects of cellular homeostasis, energy metabolism, lifespan regulation, apoptosis, DNA repair and telomere maintenance.

This resurrection of NAD importance is due in no small part to the discovery of NAD-using enzymes, especially the sirtuins. Continue reading “NAD: A Renaissance Molecule and its Role in Cell Health”

Bartonella sp. and Your Cat (or Dog, Horse, Rat): An Emerging Infectious Disease

Posted on by Kari Kenefick

If we’ve learned nothing else since February or March of 2020, we’ve learned that emerging infectious diseases are a real threat to human health globally. In a bad news/good news kind of way, Bartonellosis is an emerging infectious disease; however, it’s not spread by airborne droplets or respiration.

But if any of your family pets bring a flea or tick into the house, or if you live in proximity to mice, rats, ground squirrels, rabbits, sheep, horses or cattle–you could be at risk.

Bartonella sp. is a Gram negative, rod-shaped bacteria that has been around since ancient times. It’s the bacteria responsible for cat scratch disease (1) and for Trench fever (2), which affected soldiers during WWs I and II, and affects people living in over-crowded, unsanitary conditions around the world today.

Bartonella henselae bacteria, the causative agent of cat-scratch disease or bartonellosis,. 3D illustration
Bartonella henselae bacteria, the causative agent of cat-scratch disease or bartonellosis

Bartonella sp. are known to be spread by vectors such as fleas, which are part of the transmission cycle for cat scratch disease and the human body louse, the vector for transmission of Trench fever (3).

This animal-to-human transmission of Bartonella sp. classifies it as a zoonosis.

Infection due to Bartonella sp. often appear to be self-limiting, such as swelling in regional lymph nodes due to a cat scratch disease. In such cases, symptoms can subside without intervention. But Bartonella sp. have a nasty habit of hiding in red blood cells and in cells lining blood vessels, where they can remain undetected for a substantial period of time. This hiding place affects a host’s ability to mount an immune response, as well as affecting the ability of antibiotics to attack the bacteria.

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Sanitation and World Water Day 2021: What Would You Change about Water in Your Area?

Posted on by Kari Kenefick

Today, March 22, is World Water Day 2021, recognized by the United Nations and people around the world as a time to focus on the goal of available clean water for all.

United Nations World Water Day 2021 Graphic

Clean water for drinking is essential for our existence. A human can only survive without water for about three days.

While water is essential for life, the need goes beyond simple consumption. As is true of so many things, the COVID-19 pandemic has shown us the need for sanitation—being able to wash our hands, our clothes and ourselves with clean water, being able to rinse foods and a safe means by which to dispose of, or recycle the dirty water afterwards. And, even the need to monitor wastewater to help track infectious disease outbreaks.

World Bank and Sustainable Development Goal #6 (#SDG6)

The World Bank provides an extravagance of data on their sustainable development goal #6, Clean Water and Sanitation, noting the importance of water “for health, the environment and sustainable development”.
To add to the needs surrounding water and its scarcity, the World Bank states that:

Despite gains, more than half the world’s people lack access to safely managed sanitation services.

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Ten Things to Know about Inducible T Cell Co-stimulators (ICOS)

Posted on by Kari Kenefick

The term ICOS —inducible T cell co-stimulators— has been prominent in my work as a science writer at Promega, recently. Here is a brief look at ICOS, how it works, and how it can be used in therapeutics research and development.

T cells do amazing things, like driving or blocking production of B cells and their related antibodies and antibody maturation, and they are the primary drivers of innate immunity. T cells have a variety of surface molecules, the primary and omnipresent T cell receptor (TCR), as well as CD3.

Schematic diagram of a T cell receptor TCR. The TCR interacts with ICOS in the immune response.

In the past 15 years or so, researchers have identified other, inducible receptors on T cells. These receptors appear when T cells are stimulated, enabling interactions with other cell types. The following information is summarized from a Frontiers in Immunology review by Wikenheiser et al.

What is ICOS (inducible T cell co-stimulators)?

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