Cell Biology

Insects and Science: Optimizing Work with Sf9 Insect Cells

Posted on by Shannon Sindermann

Insects are a keystone species in the animal kingdom, often providing invaluable benefits to terrestrial ecosystems and useful services to mankind. While many of them are seen as pests (think mosquitos), others are important for pollination, waste management, and even scientific research.

Insect biotechnology, or the use of insect-derived molecules and cells to develop products, is applied in a diverse set of scientific fields including agricultural, industrial, and medical biotechnology. Insect cells have been central to many scientific advances, being utilized in recombinant protein, baculovirus, and vaccine and viral pesticide production, among other applications (5).

Therefore, as the use of insect cells becomes more widespread, understanding how they are produced, their research applications, and the scientific products that can be used with them is crucial to fostering further scientific advancements.

Primary Cell Cultures and Cell Lines

Cell culture - Cell lines - Insect Cells

In general, experimentation with individual cells, rather than full animal models, is advantageous due to improved reproducibility, decreased space requirements, less ethical concerns, and a reduction in expense. This makes primary cell cultures and cell lines essential contributors to basic scientific research.

Continue reading “Insects and Science: Optimizing Work with Sf9 Insect Cells”

2023 Promega iGEM Grant Winners: Tackling Global Problems with Synthetic Biology Solutions

Posted on by Michele Arduengo

On June 15, 2023, we announced the winners of the 2023 Promega iGEM grant. Sixty-five teams submitted applications prior to the deadline with projects ranging from creating a biosensor to detect water pollution to solving limitations for CAR-T therapy in solid tumors. The teams are asking tough questions and providing thoughtful answers as they work to tackle global problems with synthetic biology solutions. Unfortunately, we could only award nine grants. Below are summaries of the problems this year’s Promega grant winners are addressing.

UCSC iGEM

An immature night heron against the green surface of Pinto Lake. 2023 Promega iGEM Grant Winner, UCSC iGEM seeks to mitigate these harmful aglal blooms.
A night heron hunts on Pinto Lake, California.

The UCSC iGEM team from the University of California–Santa Cruz is seeking a solution to mitigate the harmful algal blooms caused by Microcystis aeruginosa in Pinto Lake, which is located in the center of a disadvantaged community and is a water source for crop irrigation. By engineering an organism to produce microcystin degrading enzymes found in certain Sphingopyxis bacteria, the goal is to reduce microcystin toxin levels in the water. The project involves isolating the genes of interest, testing their efficacy in E. coli, evaluating enzyme production and product degradation, and ultimately transforming all three genes into a single organism. The approach of in-situ enzyme production offers a potential solution without introducing modified organisms into the environment, as the enzymes naturally degrade over time.

IISc-Bengaluru

Endometriosis is a condition that affects roughly 190 million (10%) women of reproductive age worldwide. Currently, there is no treatment for endometriosis except surgery and hormonal therapy, and both approaches have limitations. The IISc-Bengaluru team at the Indian Institute of Science, Bengaluru, India, received 2023 Promega iGEM grant support to investigate the inflammatory nature of endometriosis by targeting IL-8 (interleukin-8) a cytokine. Research by other groups has snow that targeting IL-8 can reduce endometriotic tissue. This team will be attempting to create an mRNA vaccine to introduce mRNA for antibody against IL-8 into affected tissue. The team is devising a new delivery mechanism using aptides to maximize the delivery of the vaccine to the affected tissues.

Continue reading “2023 Promega iGEM Grant Winners: Tackling Global Problems with Synthetic Biology Solutions”

Genome-Wide CRISPR Screening: Putting Death on Hold

Posted on by Ken Doyle

We share this planet with approximately 8.7 million species of plants and animals. Within such a diverse environment, it’s only natural that many complex relationships have developed among different species. Some relationships are mutually beneficial, some are parasitic—and some are lethal.

Genome wide - crisper screening to help with toxic compounds to humans

Natural toxins and venoms are biologically active compounds produced by normal metabolic processes in an organism but are harmful to other organisms. Typically, toxins are encountered passively or ingested by the affected organisms, and have a specific mode of action and binding site within a cell. In contrast, venoms are introduced directly into the victim through a specialized delivery mechanism, and they may consist of a mixture of compounds that affect a range of cell types and tissues (1). Both types of poisons are produced for predation, defense, or to offer a competitive advantage (1).

Continue reading “Genome-Wide CRISPR Screening: Putting Death on Hold”

Cell Tracking Using HaloTag: Why are Scientists Chasing Cells?

Posted on by Shannon Sindermann

Cells, commonly considered the smallest unit of life, provide structure and function for all living things (3).

Eye of a fruit fly, Drosophila melanogaster, scanning electron microscopy. Scientists used HaloTag for cell tracking during eye development.
Eye of a fruit fly, Drosophila melanogaster, scanning electron microscopy

Because cells contain the fundamental molecules of life, in some situations such as yeast, a single cell can be considered the complete organism. In other situations, for more complex multicellular organisms, a multitude of cells can mature and acquire different, specialized functions (3).

Cells developing specificity are undergoing differentiation, a process where a cell’s genes are either turned “on” or “off” resultant in a more specific cell type. As these differentiated cells start to exhibit their identity, they organize themselves into the tissues, organs, and organ systems integral to the functioning of a multicellular, developing organism. This process in which order and form is created within a developing organism is referred to as morphogenesis (5).

Continue reading “Cell Tracking Using HaloTag: Why are Scientists Chasing Cells?”

Scaling Up to Measure 40,000 Data Points a Day with GloMax® Microplate Readers

Posted on by Jordan Nutting

Traditional approaches for protein degrader compound screening like Western blotting can be laborious, time consuming and cannot be streamlined with automation. By implementing a high-throughput, automated workflow that uses our CRISPER/Cas9 knock-in cell lines, live-cell bioluminescent assays and sensitive GloMax® Discover microplate readers, our custom assay services offer protein degradation profiling at an accelerated rate.  

To do this, we collaborated with HighRes® Biosolutions, to develop an automated system that can screen up to 100 384-well plates each day, generating roughly 40,000 data points with minimal hands-on work.

Learn how bioluminescent tools like HiBiT and NanoBRET™ technology can help you answer key questions in your targeted protein degradation research.

An important step of building this system is to integrate four GloMax® Discover microplate readers into the automated system using instrument’s built-in SiLA2 communication driver. The driver software makes it easy to connect the microplate readers with HighRes® Biosolution’s robotic components.

Check out our setup in the video below.

See how we’ve integrated GloMax® Discover microplate readers into a high-throughput automated system for profiling protein degraders in live cells.
Continue reading “Scaling Up to Measure 40,000 Data Points a Day with GloMax® Microplate Readers”

Our Top Three Most-Viewed Blog Posts of 2022

Posted on by Riley Bell

In 2022, our bloggers wrote on topics ranging from monkeypox outbreaks to cultured meat in biotech labs to preventing the next pandemic. Our top three most-viewed blog posts this year have the commonality of Promega products helping to advance important research in different fields and push science a step forward in the world. Take a look at Promega’s top three most-viewed blog posts of 2022.

Continue reading “Our Top Three Most-Viewed Blog Posts of 2022”

Cellular Senescence and Cancer Therapy: Overcoming Immortality?

Posted on by Ken Doyle

At the time of writing this post, no scientist had yet discovered the secret to immortality. In our world, we’ve come to accept that living things are born, grow old and die—the circle of life.

And yet, for many years, life scientists believed that the circle of life did not apply to our constituent cells when cultured in a laboratory. That is, cultured normal human cells were immortal, and they would continue to grow and proliferate forever, as long as they were provided with the necessary nutrients.

The animal cell cycle. Image by Kelvinsong; made available under the Creative Commons CC0 1.0 Universal Public Domain Dedication

Pioneering work published in 1961 by Leonard Hayflick and Paul Moorhead challenged that theory (reviewed in 1). Their research showed that normal cells in culture have a finite capacity to replicate. After they reach a certain number of replicative cycles, cells stop dividing. Hayflick and Moorhead made the important distinction between normal human cells and cultured cancer cells, which are truly immortal. In later years, the limit to the number of replicative cycles normal human cells can undergo became known as the Hayflick limit. Although some scientists still express skepticism about these findings, the Hayflick limit is widely recognized as a fundamental principle of cell biology.

Continue reading “Cellular Senescence and Cancer Therapy: Overcoming Immortality?”

Designing Better Therapeutic mAbs: An Assay for Rapid, Parallel Screening of Fc/ FcɣR Interactions

Posted on by Michele Arduengo

The first monoclonal antibody (mAb) was produced in a lab 1975, and the first therapeutic mAb was introduced in the United States to prevent kidney transplant rejection in 1986. The first mAb used in cancer treatment the anti-CD20 mAb, rituximab, was used to treat non-Hodgkin’s lymphoma and chronic lymphocytic leukemia. Today therapeutic mAbs have become a mainstay of cancer, autoimmune disease, and metabolic disease therapies and include HERCEPTIN® used to treat certain forms of breast cancer, Prolia used to treat bone loss in post-menopausal women, and Stelara used to treat autoimmune diseases like psoriatic arthritis and severe Crohn disease, among many others. Therapeutic mAbs bind targets with high specificity and affinity and they can recruit effector cells to drive target elimination through mechanisms such as antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP), making them highly specific, effective therapies.

3D rendering of a Lumit Assay which can be used for plate-based screening assay to measure the affinities of Fc interactions of therapeutic mAbs.
Continue reading “Designing Better Therapeutic mAbs: An Assay for Rapid, Parallel Screening of Fc/ FcɣR Interactions”

The Human Cell Atlas: Mapping a Cellular Landscape

Posted on by Jordan Nutting

From macrophages that seek out and destroy infectious agents to fibroblasts that hold tissues and organs together, cells give form and function to our bodies. However, despite their foundational roles in our biology, there is still much we don’t know about cells—like where different cell types are localized, what states a given cell type may take on, how the molecular characteristics of cells change over a person’s lifetime and more. Addressing these questions will provide a deeper understanding about the cellular and genetic basis of human health and disease.

Image contains several cells with a hazy outline of a DNA molecule in the background and one cell is highlighted
Continue reading “The Human Cell Atlas: Mapping a Cellular Landscape”

Cytochrome P450 Inhibition: Old Drug, New Tricks

Posted on by Darcia Schweitzer
multiwell screening plate and various pills on a table

Cytochrome P450 (CYP) inhibitors are often used as boosting agents in combination with other drugs. This drug development strategy is front and center for Paxlovid, the new anti-SARS-CoV-2 treatment from Pfizer. Paxlovid is a combination therapy, comprised of two protease inhibitors, nirmatrelvir and ritonavir. It significantly reduces the risk of COVID-19 hospitalization in high-risk adults and is ingested orally rather than injected, which is an advantage over other SARS-CoV-2 treatments, such as Remdesivir.

Nirmatrelvir was originally developed by Pfizer almost 20 years ago to treat HIV and works by blocking enzymes that help viruses replicate. Pfizer created another version of this drug to combat SARS in 2003, but, once that outbreak ended, further development was put on pause until the advent of the COVID-19 pandemic. After developing an intravenous form of nirmatrelvir early in the pandemic, Pfizer created another version that can be taken orally and combined it with ritonavir.

When ritonavir was originally developed, it wasn’t considered particularly useful because it metabolized so quickly in the body. Now it is recognized as a pharmacokinetic enhancer in combination with other drugs. Ritonivir inhibits CYP3A4, an enzyme which plays a key role in the metabolism of drugs and xenobiotics. By inhibiting CYP3A4, ritonivir slows the metabolism of other drugs. In the case of Paxlovid, this allows nirmatrelvir to stay in the body longer at a high enough concentration to be effective against the virus. This ultimately means that patients can be given lower doses of the drug with reducing efficacy.

Diagram of Nirmaltrelvir mechanism of action.
Nirmatrelvir inhibits the viral 3CL protease, so that functional, smaller viral proteins cannot be produced.
Continue reading “Cytochrome P450 Inhibition: Old Drug, New Tricks”