With global energy demand set to grow by about 47% over the next 30 years, the need for a sustainable energy transition has never been higher. Hydrogen is the most abundant element in the universe and can therefore become the perfect fuel for the future. 

Hydrogen fuel has the potential to transform multiple industries by reducing reliance on fossil fuels, providing a clean energy source, and enabling long-term energy storage. However, to make hydrogen fuel a truly sustainable solution, it must be produced efficiently and cost-effectively.

Hydrogen Future Key Takeaways

In this guide you’ll learn:

• Hydrogen fuel is a critical component of the clean energy transition.
• Advancements in green hydrogen production, fuel cell efficiency, and storage technologies are making hydrogen fuel more viable.
• Australia is at the forefront of hydrogen fuel innovation, with research institutions and companies driving significant breakthroughs.
• Precise measurement and optimisation of catalytic materials play a crucial role in improving hydrogen fuel cell performance.

What is hydrogen fuel exactly? 

Hydrogen fuel is a clean energy carrier that can be used for power generation, transportation, and industrial applications. Unlike fossil fuels, hydrogen can be burned or used in fuel cells with only water vapour as a byproduct, making it an ideal solution for reducing carbon emissions.

Normally the process of generating hydrogen requires electrical energy and if this energy comes from fossil fuels, emissions are be generated, which is not favourable. Alternatively Green hydrogen uses renewable energy to power the electrolyser that produces hydrogen from water.

Thus, if green hydrogen has to become the fuel of tomorrow, a technological breakthrough in terms of both efficiency as well as cost reduction is essential. The main cost of hydrogen fuel cells comes from expensive catalysts like platinum. 

To ensure maximum performance using the least amount of catalyst possible, it is important to carefully formulate catalytic inks for fuel cells and other applications. One key to maximising performance is the characterisation, optimisation, and control of the catalytic powder during synthesis when received, and during dispersion.

Australian Hydrogen Production as a Future Global Leader

Australia has an ambition to be a global hydrogen leader. Alongside renewable electricity, hydrogen will play a significant role in decarbonising our economy. 

The Australian government has awarded funding to multiple research projects to propel innovation in exporting renewable hydrogen to the world. Funding has been offered to research teams from nine Australian universities and research organisations. These include:  

• Australian National University
• Macquarie University
• Monash University
• Queensland University of Technology
• RMIT University 
• The University of Melbourne
• University of New South Wales
• The University of Western Australia 
• Commonwealth Scientific and Industrial Research Organisation (CSIRO)

https://arena.gov.au/news/boosting-research-into-exporting-renewable-hydrogen/

RMIT University

The Sustainable Hydrogen Energy Laboratory (SHEL) leads efforts in hydrogen production, storage, and fuel cell technologies. Key research areas include developing novel methods for hydrogen production, such as direct seawater electrolysis, and exploring efficient storage solutions. 

Notably, RMIT researchers have pioneered a technique using sound waves to enhance green hydrogen production by 14 times, offering a promising approach to affordable and sustainable hydrogen fuel. 

Hydrogen is extracted from water using sound waves which eliminates the need for corrosive electrolytes and expensive electrodes like Platinum and Iridium. 

Sound waves also prevent the build-up of hydrogen and oxygen bubbles on the electrodes, which greatly improves its conductivity and stability.

UNSW

Fuel cell is a cornerstone technology for the success of Australia’s hydrogen economy, but its scalability has been stagnant for decades because of its high cost and reliance on platinum or iridium materials. 

Researchers at UNSW are working to unlock the potential of non-precious metal catalysts for hydrogen fuel cells using an interdisciplinary approach. Highly porous, multi-site single atom catalysts will be developed to block the degradation pathways, and integrated into a novel low-water retention membrane electrode assembly. 

The expected outcomes include new materials development, new cell design and a robust platinum-free hydrogen fuel cell prototype. The project will provide significant benefits to Australia in developing revolutionary hydrogen technologies.

CSIRO

CSIRO, Australia’s national science agency, has successfully demonstrated affordable and renewable hydrogen can be generated at scale to help decarbonise heavy industry after trialling its hydrogen production technology at BlueScope’s Port Kembla Steelworks in NSW.  

Unlike conventional hydrogen electrolysers, which rely heavily on electricity to split water into hydrogen and oxygen, CSIRO’s advanced tubular solid oxide electrolysis (SOE) technology uses both waste heat (for example, steam from the steelworks) and electricity to produce hydrogen with greater efficiency.  CSIRO spinout Hadean Energy has licensed CSIRO’s SOE technology and is on a mission to accelerate industrial decarbonisation.  

RUX Energy 

RUX Energy is an advanced materials company delivering breakthrough improvements in hydrogen storage and distribution. The company manufactures patented nanoporous materials and bulk hydrogen storage systems to improve the safety, storage density and cost of hydrogen storage and distribution, making green hydrogen cost-competitive with fossil fuels.

Their Micromeritics ASAP system is providing key insight into hydrogen adsorption capacity. It enables the team to conduct quick high throughput gas sorption analysis, with the capacity to degas 12 samples and run 6 samples at once on the one instrument. 

Hysata 

Hysata based in Port Kembla NSW, is an Australian manufacturer of high efficiency electrolysers developed by a team of researchers at University of Wollongong.  This green energy start up is helping to lower costs of clean hydrogen and leading the shift away from fossil fuels to power Australian heavy industry.  Hysata has developed a ‘capillary-fed’ electrolyser that splits water into Hydrogen and oxygen. In traditional designs, gas bubbles crowd the electrode, and reduce the electrolyser efficiency. Hysata’s design uses a sponge membrane to achieve direct delivery of water to electrodes and eliminates bubble formation.

Advances in Hydrogen Fuel Cell Technology with electrolysers

What are electrolysers?

An electrolyser is a device that produces hydrogen through a chemical process (electrolysis) capable of separating the hydrogen and oxygen molecules from water using electricity. Hydrogen produced in this sustainable way, i.e. without emitting carbon dioxide into the atmosphere.

Production of electrolyser and fuel cells involves carbon supported catalyst powder, which is turned into catalytic ink and coated on a proton exchange polymer membrane. Catalytic powder contains nano-sized metal catalysts embedded in porous carbon matrix. 

Catalytic ink has a complex formulation containing Pt catalyst supported on carbon black bound by the ionomer with a range of particles and their aggregates. Some alloys, such as platinum-cobalt (PtCo), are being studied with the aim of reducing their costs by lowering the amount of precious metal required to produce fuel cells. 

Particle size, particle shape, surface area and porosity in the powder and ink plays an important role in the quality of catalyst coating in terms of homogeneity, porosity and packing density. This is another important parameter for slurry stability in terms of particle agglomeration/ sedimentation, and the amount of metal catalyst loading in the powder, ink, and coated membrane.

Optimising performance of hydrogen catalysts

The catalyst’s activity and stability are the two key parameters that determine fuel cell performance. Activity is governed by the size, dispersion, and morphology of the Pt-metal group nanoparticles. 

Equally important are the structural, textural, and surface chemistry properties of the carbonaceous agglomerates during ink drying, upon deposition on the proton exchange membrane. 

The optimised pore structure of the C-support matrix can significantly reduce the amount of Pt needed, and the optimisation of its distribution and maximisation of the availability of the catalytic points for the oxygen reduction reaction (ORR) and hydrogen-oxidation reaction (HOR) in fuel cells reduces the overall cost of the process.

Advanced Measurement and Characterisation Techniques for Hydrogen Fuel Research

Catalytic activity and stability can be optimised by developing novel Pt-alloy cathode materials, controlling the particle size for maximum mass activity, controlling the inter-crystallite distance, or ensuring uniform dispersion of Pt nanoparticles on the carbonaceous support. 

Characterisation requires a range of different particle sizing techniques such as Laser Diffraction (LD) and Dynamic light scattering (DLS) to characterise particles in different size ranges.

The Zetasizer

The Zetasizer is a DLS system that can measure the size of Carbon Black in the catalytic ink. Patented Non-Invasive Back Scatter (NIBS) technology automatically adjusts the path length according to sample characteristics like opacity and concentration. 

Thus, highly concentrated, and opaque slurries like catalytic ink can be measured delivering accurate particle size across a range of concentrations and sizes whilst maintaining consistent results. 

Additionally, Zetasizer can measure zeta potential or the charge on particles. Highly charged particles will stay dispersed while low-charged particles tend to agglomerate. See our range of Zetasizer products to find out more about the advanced light scattering system. 

Mastersizer 3000+

Mastersizer 3000+ provides another way to measure the size of carbon particles particularly when agglomerates larger than 1 µm is present in the sample. 

Mastersizer 3000+ uses laser diffraction and is considered as industry benchmark for particle sizing due to its high accuracy, repeatability and reliability. Laser diffraction is fast, non-destructive, and suited for both laboratory and continuous in-line measurements.  

It  offers a wide measurement range, from 10 nm to 3500 µm, and is well suited to cover the coarse and fine agglomerates that may be present in a catalytic powder.  While measurement of powder particles in a dry dispersion is possible, it is more common to measure them dispersed in a solvent such as isopropyl alcohol (IPA).  See our range of Mastersizer products to learn more. 

Morphologi 4 

Morphologi 4 is an automated morphological image analysis system that can be used to analyse agglomerates of the C-support particles. The Morphologi 4 can image individual particles and produce a particle size distribution based on discrete particle counting within the size range of 1 to >1,000 µm.  

Particle imaging with the Morphologi 4 is advantageous because images of individual agglomerates can be retrieved, compared, and evaluated by parameters such as circularity, convexity, and roughness. The roughness of the agglomerates is important because it could affect the ease of dispersion in the ink and the formation of porosity during deposition and drying. Check our Morphologi range to find out more. 

Micromeritics ASAP 2020

Micromeritics ASAP 2020 is a flexible gas adsorption analyser capable of measuring the hydrogen adsorption capacity of powders and porous materials. It enables the hydrogen storage capacity of new materials to be quantified which is essential for predicting the performance in a fuel cell or hydrogen storage device. 

The ASAP 2020 software has been enhanced to address the needs of fuel cell and hydrogen storage researchers. This includes: 

• Absolute pressure dosing for non-condensing probe molecules like hydrogen.
• New isotherm reports that include the weight percent of hydrogen adsorbed and the Pressure Composition Isotherm that is frequently used by hydrogen storage researchers.
• Calculated Free-space options to reduce analysis time, improve precision, and minimise exposure to interfering gases like helium.

Sample Preparation

Proper sample preparation is key for accurate hydrogen adsorption analysis, which is a two-step process. 

First, samples should be degassed on the preparation port to remove moisture and stray gases like CO₂ that absorb strongly to many materials at ambient temperature and pressure. 

Second, the sample should be degassed thoroughly on the sample port. The standard ASAP 2020 sample tube (1/2-inch stem) with a seal frit is recommended for this type of analysis. 

An isothermal jacket is recommended if the analysis is conducted at cryogenic temperatures (liquid nitrogen or liquid argon). A filler rod is optional but not recommended if the analysis is performed at cryogenic temperatures; the filler rod may interfere with the precision of low-pressure measurements. 

In summary

Laser diffraction, Dynamic Light Scattering and Automated Image Analysis and Gas Sorption can all be effective techniques for analysing various properties of pure and alloyed Pt nanoparticles supported on a C matrix. Laser diffraction is ideal for analysing the particle size of C-support that influences the Pt dispersion and porosity in the catalytic active compound. 

While morphological imaging provides some insight into the particle size, it is valuable in providing images of individual agglomerates to enable shape analysis.

Together, these techniques enable manufacturers to optimise both the cost and performance of systems based on hydrogen technology by maximising catalyst efficiency, reducing the amount of Pt required, or developing Pt-based alloy nanoparticles.

Get in Touch with ATA Scientific

If you would like to learn more about hydrogen fuel research, advanced measurement techniques, or how ATA Scientific can support your projects, please contact us today or call us on: +61 2 9541 3500

Our team is ready to assist you with innovative solutions and expert advice tailored to your research and industry needs.

References:

• https://www.dcceew.gov.au/energy/hydrogen
• https://www.malvernpanalytical.com/en/learn/knowledge-center/insights/future-days-recap-part-2-battery-and-green-hydrogen-insights-to-power-a-sustainable-future
• https://www.malvernpanalytical.com/en/industries/battery-and-energy-storage/hydrogen-catalyst-analysis
• https://micromeritics.com/resources/using-the-asap-2020-for-determining-the-hydrogen-adsorption-capacity-of-powders-and-porous-materials/
  

Quantifying the number and type of cells in a sample is a cornerstone of life sciences research, essential for ensuring reproducibility and accuracy across various disciplines, from fundamental biology to clinical diagnostics and bioproduction. Failing to count cells accurately or reliably distinguish between viable and non-viable cells can compromise research integrity, waste resources, and delay progress.

Traditionally, cell counting was performed manually, a process prone to human error, variability, and inefficiencies. However, with advancements in automated cell counting, researchers and industry professionals can now achieve greater precision, efficiency, and standardisation in their workflows.

Automated cell counting enables high-throughput analysis, reduces inter-operator variability, and provides critical data on cell concentration, viability, and morphology, optimising research outcomes and industrial applications. Whether working with mammalian cells, bacterial cultures, or plant protoplasts, adopting an automated approach ensures reliable, consistent results.

In this article, we explore the significance of automated cell counting, its advantages over traditional methods, and how innovations in fluorescence imaging and AI-powered analysis are shaping the future of cell research. You might even like to watch these webinars focused on cell counting and imaging.

The Challenges of Cell Counting

The process of cell counting starts with identifying the cells of interest, which can be quite diverse, ranging from cell lines to primary cells or even bacterial cells. If you are involved with fundamental research with cell lines or if you produce products in your facilities with cell lines, your needs will be different. Frequently, you can face the presence of debris, or contaminants, such as red blood cells if working with PBMCs or splenocytes for example. 

Also in the same sample you can have cells with different sizes, as is the case with Mesenchymal stem cells, (MCS) or you can encounter clusters of cells with different shapes in the same sample. 

You may be interested in counting other kinds of cells like pollen, protoplasts, sperm, spores, parasites  etc.. When working with bacteria extra challenges will need to be addressed including genome size and Brownian movements. 

How Automated Cell Counting Improves Research Outcomes

Counting cells is a common procedure in scientific experiments. Monitoring the health of your cells will check the proliferation rate, assess immobilisation, check the transfection rate or help you to seed cells for subsequent experiments especially cell-based assays. Therefore, it is critical to count cells using a reproducible approach especially if you wish to quantify measurements of cellular responses. 

Accurate counting provides quantitative and qualitative information such as concentration, viability, cell size, clustering, etc. Accurate counting will enable decisions to be made from passage to cell therapy development. 

In bioprocessing, where living cells are used to produce valuable products such as enzymes and monoclonal antibodies, process optimisation is critical. Accurate and consistent cell counting helps reduce inter-operator variability, a common challenge in research and production environments. 

Many applications demand total variability below 15%, and in some cases, even below 10%. Each step in the workflow introduces potential variability, underscoring the need for standardised, high-accuracy counting methods to improve experimental outcomes and enhance process efficiency.

Choosing the Right Automated Cell Counter for Your Needs

Over the past six decades, cell counting methodologies have evolved from manual techniques, counting whole cells in 2D in a Petri dish, to sophisticated automated systems, significantly enhancing efficiency and precision.

In addition to automated cell counters, the combination with fluorescent dyes can offer further insights that streamline and enhance the accuracy of nuclei assessment procedures, for example, in single cell genomics research settings. 

Fluorescent dyes used together with imaging tools such as automated cell counters, provide an effective method to evaluate nuclei quality and to quickly distinguish between living and dead cells. 

Measuring Cell Viability Using Fluorescent Dyes

The Acridine Orange (AO) and Propidium Iodide (PI) staining method offers a simple yet sensitive way to distinguish between living and dead cells. AO is a small molecule that can penetrate cell membranes of both live and dead cells and binds to nucleic acids/DNA to emit a distinctive green fluorescence.

In contrast, PI cannot penetrate live cell membranes – only membranes of compromised cells such as dying or dead cells. When PI is bound to DNA, it has peak excitation emits a distinctive red fluorescence. Using an automated fluorescence cell counter, green fluorescence of AO is observed in living cells, while red fluorescence of PI is observed in dead cells. AO/PI staining is therefore a useful and rapid technique for assessing cell viability.

The LUNA-FX7™ is a Game Changer in Automated Cell Counting

LUNA FX7™ can not only save time but also enhance research integrity by reducing errors associated with manual counting. Whether you are a researcher, lab professional, or a student counting cells for single-cell sequencing or dosing determination for a cell therapy, choosing the right cell counting technology is critical to ensuring accuracy and reliability. If you’re unsure which technology to use, contact us for a free consultation and we’d be happy to help you. 

Applications of Automated Cell Counting Across Research Fields

Human Cell Counting: Tackling Complexity with Precision

Whole blood is a notoriously challenging medium for traditional counting techniques aimed at enumerating a single component such as leukocytes. Platelets and mature red blood cells in particular complicate reliable counting. 

Similarly, CAR-T cell therapies require a complex biomanufacturing process entailing strict adherence to regulatory and QA/QC guidelines. Cell health and viability must be evaluated and monitored throughout bioprocessing workflows to ensure the safety, quality, and efficacy of the final clinical product. 

Key insights from recent case studies include:

• Leukocyte and peripheral blood mononuclear cell (PBMC) Counting: The dual fluorescent LUNA-FX7™ effectively quantifies leukocytes in whole blood using high-throughput 2-channel and 8-channel slides for replicable analyses. 

• CAR-T Cell Viability Monitoring: By employing nucleic acid stains such as Acridine Orange and Propidium Iodide (AO/PI), researchers can monitor CAR-T cell health during biomanufacturing. The LUNA-FX7™ meets rigorous QA/QC guidelines through the use of pre-set validation slides, internal QC software, and optional 21 CFR Part 11-compliant software.
• Automated QC of Isolated Nuclei:  accurate evaluation of nuclei quality is pivotal for single cell genomics research. Combining fluorescent dyes (AO/PI) with imaging tools such as automated cell counters like the LUNA-FX7™ and LUNA-FL™, provides an effective method to evaluate nuclei quality.

Animal Cell Counting: Supporting Biotechnology and Veterinary Research

Monitoring the concentration and viability of animal cells is integral to a whole hostof workflows covering areas such as livestock health, fertility assessment and cell biotechnology for protein production. 

Traditional flow cytometry-based methods can be costly and require large sample volumes, whereas the LUNA-FX7™ offers a more accessible and cost-efficient alternative.

Highlighted applications include:

• Somatic Cell Counting in Milk: Fat and protein debris complicate somatic cell count (SCC) determination, but using the LUNA-FX7™ with Somatic Cell Staining Solution enables accurate cell counting.
• SF9 Insect Cell Viability: In protein production workflows, selecting appropriate dyes and exposure settings ensures optimal SF9 cell quality assessment.
• Cattle Sperm Cell Analysis: A comparative analysis of fluorescent dyes allowed for optimised sample dilution and imaging conditions, ensuring precise fertility evaluations.

Plant Cell Counting: Immediate Viability Assessment for Protoplasts

Traditional methods for assessing the viability of plant protoplasts – plant cells with removed cell walls – require culturing until the protoplasts have developed into complete plants, making viability determination time-consuming. 

The LUNA-FX7™ overcomes this challenge by enabling immediate assessment using double-staining with distinct-coloured fluorescent dyes. Researchers have successfully identified optimal dye combinations and parameters, allowing for reliable and rapid viability assessment, streamlining workflows in plant biotechnology and genetic engineering.

Innovations in Automated Cell Counting: Embracing AI and Machine Learning

Cell counting remains an essential process across fundamental research, clinical applications and bioproduction. As science continues to advance, there is a growing demand for faster, higher-throughput, and more tailored cell counting solutions. While automated techniques address many of these demands, challenges including sample prep, technology compatibility, cost efficiency, and compliance remain. 

Recent innovations in software and hardware, including the integration of artificial intelligence and machine learning, focus on enhancing accuracy and efficiency in cell counting. For instance, the new LUNA-III™ cell counter utilises machine learning algorithms to ensure high-quality and reliable results, addressing current cell counting challenges and providing practical solutions for applications such as CAR-T cell therapy production and single-cell sequencing.

Selecting the appropriate cell counting technology is crucial for research success. Factors to consider include sample preparation requirements, technology compatibility, cost efficiency, and compliance with regulatory standards. 

Automated systems like the LUNA-FX7™ and LUNA-III™ offer advanced features that cater to these considerations, providing researchers with tools to enhance accuracy, efficiency, and reproducibility in their workflows.

Are you ready to upgrade your lab’s cell counting capabilities? Explore how the LUNA-FX7™ and LUNA-III™ can transform your workflows and enhance research outcomes today!

Contact us for a demo!

Reference: SelectScience releases exclusive Application eBook for cell counting accuracy and efficiency – Logos Biosystems | Advanced Imaging Solutions for Research Excellence

“This is where formulations come to die!” These are words of a prominent pharmaceutical company GMP formulations lead. What makes this so difficult? How can a formulation go this far before falling over? Presumably there have been extensive clinical trials, yet as soon as scaling is called for something fails. What makes a winning formulation? This article will explore some key points to consider when formulating medicinal products, predominantly for in vivo injection. 

Many years ago, at a conference in Chicago, a pharmaceutical company representative asked advice on how to analyse a tablet post digestion – because they ‘had no idea what really happens to their oral medications once swallowed’. This sent me into a spin… Since then, I have sought alternatives for drug delivery.

Intravenous (IV), subcutaneous (SC), and Intramuscular (IM) routes help reduce the actives’ concentration requirement plus preserves the payload from the oral degradation. They enjoy a rapid onset of action, are apparently a predictable way of action and almost complete bioavailability. Given the nature of the payloads considered, this chapter will focus on routes other than oral.

How do we encapsulate nanomedicines for targeted delivery

Common reasons to encapsulate Protein, peptide, RNA, or small molecules are many, such as to protect the payload and to facilitate cellular uptake. Other proposed uses include the incorporation of a ‘decoration’ to guide the nanoparticle to a particular target, though there is little evidence of this being very specific. Novel nanoparticles looking at changes in structure and constituents are increasingly being employed. Importantly a nanocarrier will reduce systemic toxicity, reduce concentrations of the payload required, and protect a payload from premature biological degradation. 

How can we currently encapsulate?

Over the years there has been a dramatic increase in microfluidic devices from an array of companies attempting to cash in on the explosion of research, particularly with RNA of one flavour or another.
I think it was Harry Truman who stated, “If you can’t convince them, confuse them”, I aim to demystify this expanding world and hopefully bring clarity to this confusion.  If you invest time in reading the white paper https://www.atascientific.com.au/a-better-way-to-create-nanoparticles/ you will find a discussion about an array of microfluidic devices, how they work and the limit of their uses. To expand, this document relates to the formulation itself and its interaction with mixer employed. 

Formulation as a function of prevailing technology

Many companies and research institutes invest a deal of resources formulating a lipid construct that will not only encapsulate the required cargo but mix well to elicit the targeted size and Poly Dispersity Index. This is the current paradigm. Unfortunately, each mixer has a peculiar method where, at times, the formulation fails with orthogonal technologies or fails to scale efficiently. Some organisations invest in low-cost T Mixers or similar only to learn that they need to spend enormous human resources to develop a formulation to efficiently produce poultry volumes. In the Australian parlance, this is an apt descriptor for “the tail wagging the dog”. Ideally the most versatile technology would be one that has the capacity to morph to a formulation. 

If I had a winning formulation, I would be reticent to modify it for a mixer. I recall a conversation I had with the Director of CMC in a Canadian therapeutics’ organisation, he stated “We make exceptional formulations; You just have to mix it”. Whilst it sounds flippant, he was prompt to add, the technology we presented was adept at transforming to suit any given formulation, indeed in “two runs” we had nailed a formulation they had to tweak for 6 months on another technology.

It is becoming increasingly clear that formulations may be stymied by the technology employed. The inflexibility of many microfluidic devices coupled with the abject failure to scale is concerning. The lure of an easy fix or popular choice of peers may in fact limit the very research you wish to expand, not only in terms of expense, time, and resource but opportunity. Many formulations are likely to have failed as they approached the limit capacity of the employed mixer.  Following the golden rules of nanoparticle creation- mixing rate must be faster than assembly rate.  

Nanoprecipitation can be tuned with changes to flow rate and the flow rate ratio of the two phases mixed. Well, this is the case with most instruments, what if you can make minor modifications of the technology to enhance the creation of bespoke formulations, potentially turning what was thought to be marginal into a lead candidate. In the discussion below about PEG alternatives, I pondered their acceptable results and wondered what if these were a function of the technology limit- not the lipid’s limit? Frequently I have run formulations where microfluidics yield Encapsulation Efficiency (EE) of around 80– 90% – acceptable? Not really as I approach 99 – 100% on the exact same formulation. 

Quality of nanoparticles

Assessing the apparent quality of a nanoparticle is interesting! To what measure or standard?
The current theory starts with Dynamic Light Scattering with the gold standard system – the Malvern Panalytical Zetasizer, analysing the Size, Poly Dispersity Index (PDI) and ZetaPotential. The next question is the Encapsulation Efficiency (EE) a measure of how much of the payload eg API or RNA is actually held within the construct of the nanoparticle. Ideally it is as close to 100% as possible as the cost of some payloads are significant, where every 1% reduction in EE will have dramatic effects on total costs of production. – if you are not encapsulating you are virtually flushing away the expensive RNA. There needs to be more research into analytics of the payload particularly what is the effect on the payload once it is encapsulated into a nanoparticle? Such needs are heightened should multiple APIs be encapsulated within the same nanoparticle, what interactions occur? Some interesting research into this arena with RNA is occurring with the advent of Microfluidic Modulation Spectroscopy, a technology capable of elucidating protein structure. Recently an enhancement enabled easy detection and comparison of RNA changes that are vital for therapeutic development, and here is the kicker – within an LNP.

Checking the in-vivo capabilities of the particle is where ‘the rubber hits the road’. Understanding transfection, bio-distribution and efficacy is the next level of assessment. Clearly this will define the effectiveness of the delivery vehicle and the payload as well. A long-held debate has been whether the murine model is a valid measure. A recent paper from the University of British Columbia (UBC) has some interesting findings with a porcine model¹, demonstrating “… that following a low-dose infusion of mRNA-LNP, exogenous protein and exogenous mRNA transcripts can be detected globally including in the liver, spleen, lung, heart, uterus, colon, stomach, kidney, small intestine, and brain of the swine. Notably, we also detected exogenous protein expression in the bone marrow, including megakaryocytes, hematopoietic stem cells (HSC), granulocytes, and in circulating blood cells such as white blood cells and platelets.”¹ It has, in the past, been widely accepted to attain extrahepatic targeting employment of a specialised moiety decorating the surface of a Lipid NanoParticle (LNP) is required, perhaps this may not be a necessity. 

Not all Lipids are equal

The complexity of formulation does not end with the achievement of appropriate particle size, PDI, zetapotential and EE. Even with an EE of 99% the question remains whether there is adequate cellular uptake, does the payload release where intended, is it taken up by the intracellular organelles targeted, what happens in endosomal escape? Most of these are poorly understood. Screening of a lipid candidate can include biodistribution studies where expression of a fluorescent marker is tracked by imaging, or alternate methods. Perhaps it is the formation of antibodies that define success. “Despite the developments in the LNP field, a commonly overlooked aspect regards the very limited release of the nucleic acid payloads in the cytoplasm¹⁰.”

“The highly dynamic nature of endosomal compartments and shared markers between them adds another layer of complexity making it even more difficult to gain common conclusion regarding trafficking and escape of LNPs. Nevertheless, endosomal escape is one of the most important aspects for efficient nucleic acid delivery, and it is also pertinent to pay attention to the requirement of sophisticated tools and probes to understand the basic molecular biology of this process¹⁰.”  

Lipids may not be equal, neither are the payloads and their purpose. It has been noted that there are many lipids that combine to form an LNP, as Robin Shattock’s group¹¹ reports, the helper lipid selection can have a significant role.  They noted ‘helper lipid identity altered saRNA expression’ and ‘LNP storage’. Confirming formulation is a richly diverse field with a great deal to contribute to medicine. 

It may not be just about vaccination or chasing a tumour, it may be as fundamental as enabling better platelet transfusions. Exciting research at UBC has succeeded in transfecting platelets with mRNA via an LNP, to ‘enable exogenous protein expression in human and rat platelets’¹². It is thought to be the first instance of this and could possibly expand the therapeutic potential of platelets. 

Alternate Formulations

Metal-phenolic networks (MPN) are offering another indication there could be alternatives to the current paradigm. The Caruso lab in Melbourne University have developed a method to create MPN Nanoparticles² in not only a straightforward reproducible, but tunable manner. “We demonstrate the role of buffers (e.g., phosphate buffer) in governing NP formation and the engineering of the NP physicochemical properties (e.g., tunable sizes from 50 to 270 nm) by altering the assembly conditions. A library of MPN NPs is prepared using natural polyphenols and various metal ions. Diverse functional cargos, including anticancer drugs and proteins with different molecular weights and isoelectric points, are readily loaded within the NPs for various applications (e.g., biocatalysis, therapeutic delivery) by direct mixing, without surface modification, owing to the strong affinity of polyphenols to various guest molecules.²” This is hugely impactful work with global implications. Well worth following this lab for the exciting things to come. 

Polysarcosine lipids – PEG alternative

Lipid nanoparticles (LNPs) such as ALC-0315 and SM-102 LNPs have been applied to deliver mRNAs encoding viral antigens against the SARS-CoV-2 virus. These LNPs are typically comprised of four distinct lipid components: ionizable lipids, phospholipids, cholesterol, and polyethylene glycol lipids (PEG lipids)³. Prior to the COVID Pandemic there were studies evaluating the prevalence of pre-existing anti-PEG antibodies⁴. The use of this lipid construct across the globe to vaccinate billions of people prompted over a dozen studies regarding anti-PEG antibodies occurrence, defining which immunoglobulin was triggered and resultant allergic responses. As reported by Kang³ the results were conflicting. Despite the inconclusive nature of these studies, there is a growing appeal for alternatives to PEG.  Kang³ evaluated a panel of polysarcosine (pSar) as a direct replacement to PEG in the ALC-0315 and SM-102 based LNP formulations with comparable physicochemical properties perhaps even enhancing mRNA delivery efficiency in-vitro and in-vivo along with overall immunogenicity. Interesting developments that may assist all LNPs. 

Poly-lactic-co-glycolic acid (PLGA)

PLGA are a family of FDA-approved biodegradable polymers that are physically strong and highly biocompatible and have been extensively studied as delivery vehicles of drugs, proteins, and macromolecules such as DNA and RNA⁶. Over the years there have been few more enduring materials in drug delivery research with such a wide range of applicability than that of Poly-lactic-co-glycolic acid (PLGA). Such a wonderful biodegradable polymer that essentially breaks down to Lactic and Galactic acids and ultimately CO₂ and H₂O eliminated by the human body through a natural pathway such as the Krebs cycle. As diverse as the applications, so are the methods to create PLGA particles, such as Spray drying, Microfluidics and nanoprecipitation with lipids to name just a few. Most methods struggle to create enviable polydispersity and volumetric yield. The methods are laborious and difficult to scale. A solution to this is the Advanced Cross Flow (AXF)⁵ where Membrane emulsification operates in a controlled low shear environment, bringing the advantages of microfluidic mixing to the commercial scale. By substantially reducing the energy input membrane emulsification dramatically improves particle size distribution when compared to traditional batch homogenization. The result is a more consistent product that is not lost to downstream filtration. 

Moreover, AXF technology operates continuously, enabling real time process monitoring and feedback control during drug product manufacturing. A simple change to the membrane tunes the system to produce either nano or Micron sized particles. One installed system has been configured to create 10 Kg of PLGA particles per day, dispelling the fear of lack of yield. In a search Aug 2024, of ClinicalTrials.gov (https://clinicaltrials.gov/search?term=PLGA&intr=PLGA ) there were 43 with PLGA particles.

Freeze dried lyophilised 

Whilst not strictly a novel formulation method, freeze drying is interesting as a delivery vehicle for other formulations. Powdered forms of nanoparticles could be highly efficient nasal delivery systems.
“The lung is directly exposed to the outside environment through the airways. It contains two main functional parts, the conducting zone (trachea, bronchi and bronchioles) and respiratory zone (alveoli). The top five most common lung diseases causing severe illness and death worldwide include tuberculosis, respiratory infections, lung cancer, asthma and chronic obstructive pulmonary disease (COPD), which together have a huge global burden⁷.” Imagine the global health significance of a loaded LNP, freeze-dried and delivered by inhaler direct to the lungs. Sending anything into the lungs is not as simple as it sounds. Consider that at its core, the lung is a site for gaseous exchange, but unfettered access would expose the body to innumerable airborne pathogens.

 Evolution has armed us with some formidable defenses such as a wash of antimicrobials, mucus, neutralising immunoglobulins, beating cilia and an epithelial cell layer. These are bolstered by white blood cells. This is why packaging the drug to move by stealth enhances the success of treatment.

Liquid Crystalline Nanoparticles (LCNP)

Distinctive structural features of lyotropic nonlamellar liquid crystalline nanoparticles (LCNPs), such as Cubosomes and Hexosomes enhance their applicability as effective drug delivery systems. “Cubosomes have a lipid bilayer that makes a membrane lattice with two water channels that are intertwined. Hexosomes are inverse hexagonal phases made of an infinite number of hexagonal lattices that are tightly connected with water channels. These nanostructures are often stabilized by surfactants. The structure’s membrane has a much larger surface area than that of other lipid nanoparticles, which makes it possible to load therapeutic molecules. In addition, the composition of mesophases can be modified by pore diameters, thus influencing drug release⁸”. This method has the potential for a kind of repackaging of existing medications for more efficient and effective delivery.
In context- antimicrobial drugs may be able to pass into cells, therefore the previously unreachable intracellular bacteria can be treated. Consider chronic lung infections. An example of how effective this is was the subject of a study at the University of South Australia, ´The ability of a cationic LCNP to improve the performance of antibiotics against intracellular bacteria present in macrophage and epithelial cells was determined. The presence of DDAB altered the morphology of LCNPs into an onion-ring like structure. The inclusion of a cationic lipid enhanced cellular uptake, >50% and > 90% in macrophages and epithelial cells, respectively. In macrophages, LCNP-DDAB reduced intracellular P. aeruginosa and S. aureus viability by ∼ 90% and ∼ 55% at 4x MIC of antibiotics.⁹’

Looking toward the future of nanoparticle formulation

Will there be one magic bullet or horses for courses approach? If history boasts a rich tapestry of methods for drug delivery, the future is sure to be exciting. This review is not definitive, it is moreover a snapshot of a few prevalent procedures in a constantly evolving environment.

The creation of nanoparticles has nuances, being able to pivot to these with a flexible technology will likely be the successful combination going forward. Microfluidic devices have challenges when called upon to scale up despite the promise of a limited interventionary technique. Their failure has prompted a paradigm shift to a novel method of nanoparticle production, the Micropore Technologies Advanced Cross Flow systems are a solution with ample flexibility to take you from 200 ul R&D samples to thousands of litres in GMP production using the same technology in R&D as in GMP without being formulation centric.  Whilst not a panacea for all delivery goals, this technology sure has legs for the lions’ share.

References

1. ´Protein is expressed in all major organs after intravenous infusion of mRNA-lipid nanoparticles in swine’ Ferraresso et al 2024 Molecular Therapy: Methods & Clinical Development https://doi.org/10.1016/j.omtm.2024.101314
2. ‘Direct Assembly of Metal-Phenolic Network Nanoparticles for Biomedical Applications’ Xu et al. Angewandte Chemie International Edition https://doi.org/10.1002/anie.202312925
3. ‘Engineering LNPs with polysarcosine lipids for mRNA delivery’ Kang Et Al, Bioactive Materials 37 (2024) 86-93. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10957522/pdf/main.pdf
4. Analysis of Pre-existing IgG and IgM Antibodies against Polyethylene Glycol (PEG) in the General Population Yang . Q. Et Al Anal Chem. 2016 Dec 6;88(23):11804-11812.
   doi: 10.1021/acs.analchem.6b03437. Epub 2016 Nov 16.
5. ‘Polymeric Encapsulation of Active Pharmaceutical Ingredients’ https://microporetech.com/applications/biodegradable-polymeric-materials
6. ‘PLGA-Based Nanomedicine: History of Advancement and Development in Clinical Applications of Multiple Diseases’ Alsaabet Al, Pharmaceutics. 2022 Dec; 14(12): 2728 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9786338/
7. ‘Lipid Nanoparticles as Delivery Vehicles for Inhaled Therapeutics’ Leong et al Biomedicines. 2022 Sep; 10(9): 2179. doi: 10.3390/biomedicines10092179
8. ‘Recent Advances in the Development of Liquid Crystalline Nanoparticles as Drug Delivery Systems’ Leu et al Pharmaceutics. 2023 May; 15(5): 1421.
   doi: 10.3390/pharmaceutics15051421
9. ‘Liquid crystalline lipid nanoparticles improve the antibacterial activity of tobramycin and vancomycin against intracellular Pseudomonas aeruginosa and Staphylococcus aureus’ Subramaniam et al International Journal of Pharmaceutics Volume 639, 25 May 2023, 122927, https://www.sciencedirect.com/science/article/pii/S0378517323003472#b0240
10.  ‘Endosomal escape: A bottleneck for LNP-mediated therapeutics’ Chatterjee et al; PNAS March 2024: https://doi.org/10.1073/pnas.2307800120
11. ‘The role of helper lipids in optimising nanoparticle formulations of self-amplifying RNA´ Barbieri et al; Journal of Controlled Release Volume 374 , October 2024, Pages 280-292 https://doi.org/10.1016/j.jconrel.2024.08.016
12. ‘Genetically engineered transfusable platelets using mRNA lipid nanoparticles’ Leung et al;
    Sci. Adv. 9, eadi0508 (2023), Downloaded from https://www.science.org on September 02, 2024

Ease of scalability

Producing COVID-19 vaccines for the entire world population rapidly demonstrated the microfluidic development scale up roadblock. Discovery formulations developed using microfluidics had to be scaled up using alternative higher throughput formulating technology with different operating parameters.   

With a growing demand for more mRNA-LNP based vaccines, proven to have high efficacy, there is currently a need to facilitate a faster and cheaper global deployment of a high-throughput manufacturing method. A GMP compliant approach is needed, from the earliest possible stage through to manufacturing, that is also tuneable in size to access various tissues or specific drug targets.  

Advanced cross flow mixing (AXF) from Micropore Technologies allows for seamless scaleup with consistent physics, mechanisms conditions and geometry across its equipment range. Ultimately Micropores AXF technology can be scaled up to a device with over 10 million pores with the potential for a throughput of up to 20 Liters per minute all this from a device that would still fit inside a briefcase. AXF allows controlled, low shear precision continuous flow mixing technology from nano to micro formulations for when you want to avoid roadblocks on your product development journey. It uses the same same shear, same physics and same technology from lab bench to manufacturing scale to enable scale up with confidence.  

Micropore technology address key concerns for adoption

• Expertise, availability of skilled personnel, associated risks, and the need for a reliable supply chain. 

Micropore is a technology provider with global experience in manufacturing all different types of vaccine modalities can further ensure a cost-effective, high-quality process. Partnering with Micropore will enable a stronger benchmark with in-depth expertise and the ability to leverage novel technologies will also help reduce risk and shorten timelines.

• There is a need to develop standard analytical methods for quality characterisation and reference material that lend insight into the mechanisms of stability or degradation of mRNA and LNPs containing it. 

The determinants of stability of mRNA in LNP formulations – what parts are predicated on the payload of mRNA and what portions are predicated on the lipid nanoparticles themselves or what portions are predicated on the freeze-drying cycle that’s used – can be related to its size and its secondary structure. Messenger RNA poses a unique manufacturing challenge because of its large size. Other RNA entities such as siRNA and guide RNA for clustered regularly interspaced short palindromic repeats (CRISPR) technology typically are produced using chemical synthesis, which can be performed in a relatively controlled environment. But mRNAs are larger, with complex three dimensional structures that aren’t yet fully understood. Malvern Zetasizer (DLS) enables particle size and stability measurements while the RedShiftBio Aurora (MMS) system enables secondary structure (HOS) determination.

• The need to move to larger scale manufacturing of mRNA-LNPs without an extensive setup or excess capacity to convert.

With mRNA vaccine production requiring relatively less space than other approaches, new facilities may be more feasible and affordable. Micropore technology can enable localised production of vaccines and thus accelerate access to a much larger population. In locations with limited or no infrastructure, the Micropore approach can be the shortest route to production and can reflect the exact needs of the organisation at minimal cost.

• The need for future vaccine manufacturing facilities to be closed and continuous processing, plus equipment connectivity and communication.

Micropore offers a minimal cost model achieved through the AXF advanced cross flow technology platform. The flexibility of mRNA-based vaccines when manufactured using this single piece of stainless steel equipment with no consumables means it requires the least capital investment. The scalability of production (from 200 μL to 1500 L/hr) reduces the facility design complexity and means that more doses can be manufactured in a continuous process which eliminates variability, compared to a batch-to-batch approach. As such, this vaccine modality combined with the Micropore mixing platform can be a robust starting point for production with low risk.

• The need to overcome current barriers of manufacturing process to be fully digitalised as regulatory authorities rely on data and parameters recorded during production for verification and approval.

Micropore cross flow technology demonstrates predicable scalability which is favourable especially for GMP manufacturing which means process controls can be introduced that are automated – process analytical technologies (PAT). Automated analysis of properties such as online particle size enables the option to automatically control any deviations in size and feed that back to control the pumps and optimise control to give the correct size again. This increases confidence in quality of production meaning throughput can be increased further.

Continuous Formation & Stabilisation of LNPs Minimises Risk of mRNA Degradation

Batch mode vs continuous mode manufacturing: While Impingement jet mixing (IJM) or T-mixers are the most widespread manufacturing method currently, high turbulent mixing combined with high pressure and shear stress effects can compromise LNP stability and affect overall performance of the product. The process also suffers from high batch variability and high wastage due to holding or prolonged processing steps.

Micropore Technologies offers Continuous Manufacturing: Micropore Technologies employs laminar flow mixing across a permanent stainless-steel membrane to produce reproducible, scalable LNPs. The outer dispersed phase is continuously mixed with the aqueous inner compartment to form LNPs. Size controlled uniform particles are generated in a continuous flow capacity of up to 1500 L/hour making this by far the fastest production rate in the LNP industry. This would translate to roughly 58,000 doses of vaccine every minute, an important capability when faced with the demands of global disease emergencies. 

Micropore Technologies: 2^nd generation Vaccine manufacturing Process

Micropore’s Membrane Technology: How it works

Micropore’s equipment is uniquely suited to the production of complex nanomedicines in a scalable manner – from small 200µl samples through to 1 billion doses.

Micropore technologies differs from conventional mixing LNP techniques by considering the process not as discrete or separate unit operations but as one single whole process.

Attributes:

• High efficiency, scalable and reproducible.

• Reduced manufacturing time and costs.

• Customisation for diverse LNP formulations.

• Tunable particle size with narrow size distribution (PDI).

The Micropore system comprises a membrane inside a housing. Lipid mixtures added from the top are mixed with the RNA/buffer solution from the side which enters the membrane to produce monodisperse particles.  The system operates at very low pressure – approximately 2 Bar. This allows the size of the particles created to be very predictable with any change in flow rate. T-mixer devices operate at much higher pressures – about 30 Bar.

The graphs above show comparison control curves for AXF vs. T-mixer.

Advanced Cross Flow (AXF) mixing

Micropore’s equipment is uniquely suited to the production of complex nanomedicines in a scalable manner – from mL batches up to tonnes.

A lipid/organic phase passes through the 100,000s of membrane pores into the flow of aqueous continuous phase that passes through the centre of the membrane tube. Gentle, laminar flow based mixing allows for good preservation of sensitive materials. Precision engineered equipment gives great size distributions even at scale, allowing precise targeting of distributions.

Process robustness and high encapsulation efficiency

Here we present data from a formulation prepared using the AXF Pathfinder 20.

The conditions used are listed below:

• Formulation ratio DDAB:DSPC:cholesterol:DMS-PEG2000, 40:10:48:2 mol%.
• Lipid concentration, 3.5mg/ml
• N:P ratio, 1:6
• polyA concentration, 0.046mg/ml
• Flow rate ratio 3:1
• Total flow rate 100mL/min
• continuous phase, Tris buffer, 10mM pH 7.4

The Pathfinder delivered highly repeatable, monodisperse LNPs that were 70nm in size with a narrow PDI of 0.13. The experiment was repeated in triplicate and samples were measured using the Malvern Zetasizer DLS system.

Encapsulation efficiency is about 97-98%. The AXF mini uses cross flow mixing which is a gentle mixing technique and prevents RNA degradation.

Process tunability and predictability

The two graphs above demonstrate the superior capability of the AXF mini to control the size of nanoparticles produced which can be critically important, especially for GMP manufacturing.

The plot on the right shows two formulations, plotted in red and green. Differences arise from e.g. concentration of DMG-PEG200 in the range 0.004 – 0.12 μmol which reduces size from 200 nm to 30 nm but both exhibit the same shape and therefore control. Although the PDI is still quite tight in both formulations but the formulation plotted in green is considerably larger (100-160nm) compared to the formulation in red (50-110nm). From this experiment you can quickly see it is possible to produce a 70nm particle by using a total flow rate around 100 ml/min. 

The plot on the left shows an experiment where the AXF mini was used to create two different formulations using two different flow rates. At 20ml/min the LNP size was 107 nm. When the total flow rate was increased to 200 ml/min, the LNP size reduced to 55 nm with a remarkably low PDI of 0.06.  

This data demonstrates the predicable scalability which is favourable especially for GMP manufacturing and regulators which means you can start to introduce process controls that are automated – process analytical technologies (PAT). Automated analysis of properties such as online particle size enables the option to automatically control any deviations in size and feed that back to control the pumps and optimise control and give the correct size again. This increases confidence in quality of production meaning throughput can be increased further. ATA Scientific offers several technologies to characterise LNPs.

Contact us for a demo today!

Live-cell imaging is important for many applications however limitations of conventional methods have constrained its routine use. Reliable live cell imaging requires an environment that keeps the cells functioning during the experiment while also being able to ensure the experimental method is not perturbing the cells and affecting the interpretation of the results. Here we discuss the growing popularity of automated live-cell imaging systems and highlight some key features to look for when selecting a live cell imaging system.

What is live-cell imaging?

Live-cell imaging is a microscopy-based technique used to examine living cells in real-time. It offers deeper insights into dynamic cellular processes such as migration, confluency and signaling and can reveal findings that might otherwise have been overlooked. Both brightfield and fluorescence-based live-cell imaging modalities support a range of different analysis needs.

How is live-cell imaging used?

Applications of live-cell imaging span basic research through to biopharmaceutical manufacturing. In a research setting, live-cell imaging can be used during cell culture to help define the best time for harvest, determine the senescence status of cells, assess drug treatments for cytotoxicity or detect and monitor phagocytosis. For example, Phagocytosis, is a process by which certain live cells, called phagocytes internalise foreign matter. This defensive reaction against infection is key in the study of immunology and plays an important role in immune responses, tissue homeostasis, and continuous clearance of apoptotic cells. Generally, phagocytotic activity is assayed using flow cytometry. However, this process only provides quantitative data and does not provide the means to monitor phagocytosis in real time. Performing fluorescence-based assays using the CELENA® X High Content Imaging System with pH-sensitive fluorescent particles, like pHrodo™ Green, can be an effective and efficient system for quantifying and monitoring apoptosis activity. For biopharmaceutical manufacturing, live-cell imaging has broad utility for process development and control throughout the production of biologic drugs and vaccines.

Limitations of conventional live-cell imaging methods?

Historically, live-cell imaging has involved manually monitoring cells by culturing them in a CO2 incubator. The culture vessel is removed several times to take images of cells over time using a digital microscope. This approach is labor-intensive and highly prone to human error, largely because it offers no means of finding the same position in the culture vessel. Fluctuating environmental conditions can also cause cellular stresses, which can compromise results. While benchtop imaging systems improve on this method, they are bulky and cumbersome, and often struggle to maintain a stable environment.

3 Factors to consider when choosing an automated live-cell imaging system

Automated live-cell imaging systems like the CELENA® X offer a flexible design that is smaller, faster and easier to use to meet both the demands of the drug discovery industry and the basic research needs of the smaller laboratory.

Multiple imaging modes that are affordable

Live cell imaging systems that offer both brightfield and fluorescence options for either time-lapse or real-time monitoring offer maximum flexibility. Celena X integrates an automated fluorescence microscope with quantitative image analysis software to process large datasets at an affordable cost. Its interface allows a user to run multi-well or multi-spectral experiments with capacity for multi-point imaging with only a few clicks. The microscope provides for a multitude of fluorescence cell imaging possibilities by supporting all objective magnification from 1.25x to 100x, both brightfield and phase contrast illumination, and with LED filter cubes.

Stable scanning performance and compatibility

The system is compatible with a wide range of cell culture vessels such as multi-well plates, dishes, flasks and slides to cover a wide variety of assay types. Depending upon the application, either image-based or laser-based autofocus methods can be used. The CELENA® X can be used for image confluency in McCoy cells seeded in 96-well plates over 48 hours with brightfield image-based autofocusing, demonstrating how the system can be modified and applied as a high throughput method for various cell-based assays. With laser-based autofocusing and multiple filter cubes, this method utilised the CELENA® X to image the dose-dependent effects of anti-cancer drugs throughout the cell cycle in HeLa cells seeded in 96-well plates, demonstrating how the system can be used in a multivariate drug screening process.

User friendly interface and 3D modeling

An analysis of high content images with large datasets can cause problems with most types of analytical software. Each new assay requires the creation of new modules, which can be challenging for lab staff and often involves IT staff. The CELENA® X provides an easy-to-use modular-design analysis software based on a powerful CellProfiler engine. Tens of thousands of images can be analysed automatically to obtain quantitative information with a complex software setting or optimisation.

Three-dimensional (3D) cell models can provide a more accurate representation of real cell environments than 2D cell culture systems but requires a different strategy of imaging and analysis compared to 2D cell culture. Organoids, for example, are organ-specific 3D cell models derived from human stem cells, designed to mimic the functionality and structure of human organs while 3D spheroids can represent a gradient of nutrients and oxygen between cells located in both outer and inner layers which is more relevant to physiological environments. These 3D models are notably useful for studying various types of cancers.

The challenge when imaging organoid and spheroid assays comes from organoids having multiple focal planes making it difficult to acquire in-focused images for multiple organoids. For live/dead cell viability of the single organoid, a different analysis strategy is required since individual cells in an organoid do not exist as a single live/dead status. To address this issue, CELENA X employs MergeFocus software module after acquiring Z-stack images from multi-channel fluorescence.

We invite you to use the Celena X automated live cell imaging system today and compare it for yourself.

Contact us to arrange a free trial.

ATA Scientific Pty Ltd
+61 2 9541 3500
enquiries@atascientific.com.au 
www.atascientific.com.au 

Product page link: – https://www.atascientific.com.au/products/celena-x-high-content-auto-cell-imaging-system/