Viewpoint article on engineering cell surfaces published in ACS Macroletters
Engineering cell surfaces is not trivial. In Nature gene regulation controls protein expression, but we are interested in non-genetic tools to introduce new functionality to cell surfaces, specifically synthetic polymers. In this viewpoint we summarize how the technique of metabolic glycan labeling is being used in polymer/nanoscience to introduce non-natural functionality to cell surfaces. Metabolic glycan labeling allows the introduction of bio-orthogonal handles (e.g. azides) to cells by hijacking glycan biosynthesis pathways by addition of e.g. Azido-N-acetyl mannosamine derivatives, which is processed into sialic acid on the cell surface. This azide (or other functionality) can be used to recruit polymers to the cell surface by formation of a covalent bond. In this article we highlight this really exciting area of biomaterials and summarises some applications including cell-tracking, studying cell-cell communication and more.
This article is part of the 100th Anniversary of Macromolecular Science series.
Our latest work on macromolecular cryoprotectants has been published in Biomacromolecules
There is increasing interest in developing new macromolecular cryoprotectants to improve cryopreservation outcomes, however, the criteria used for assessing cryopreservation success varies greatly between studies. In this work we critically analysed the impact of different macromolecular cryoprotectants on post-thaw cell viability and cell recovery, at multiple timepoints, for several cell lines. We found that cell viability was not a good predictor of cryopreservation outcome as it overlooked the number of cells lost during the cryopreservation process. In addition, we showed that it is essential to culture the cells for a period after thawing to allow apoptosis (programmed cell death) to initiate and complete. Considering these findings, we demonstrated that polyampholytes (an emerging class of macromolecular cryoprotectant) are effective cryoprotectants that improve both cell viability and cell recovery, compared to poly(ethylene glycol) which can produce in false positive results.
Read the paper here:
Glyconanomaterials to probe influenza Hemagglutinins is published in Biomacromolecules
Pathogens invade their hosts by several mechanisms including binding to glycans (sugars) on our cell surface. This binding is a crucial stage in their infection cycle and understanding these processes and developing new diagnostics and treatments. In this work we used glycoyslated nanoparticles to investigate how carbohydrate-binding proteins on the surface of influenza (hemagglutinins) bind sialic acids. We used our gold nanoparticle platform to enable easy incorporation of the sialic acids at the ends of polymers immobilised onto the gold particles, ensuring they were colloidally stable but still capable of present the sialic acids. Using a range of assays we optimized their structure to maximise outputs. We then interrogated a panel of hemaglutinins from different influenza strains, including zoonotic (species-crossing) strains. Crucially, when influenza 'jumps species' (e.g. avian to human), the nature of the glycan it binds also changes, which we were able to rapidly map. These results showed that our nanoparticle platform is a suitable tool for interrogating viral surface proteins and for helping to understand zoonosis.
Latest work published in Materials Horizons exploits new polymeric nanomaterials to modulate ice growth.
Polymerisation-induced self-assembly (PISA), a scalable and versatile method to obtain soft nano-objects is utilised for the first time to introduce ice recrystallisation inhibiting (IRI) polymer nanomaterials. Crucially we developed a method to ensure the particles are saline stable which is essential for IRI testing but current PISA formulations cannot tolerate any salt. Uniquely, we achieved this by tuning the core, enabling us to retain our IRI active corona (based on poly(vinyl alcohol)). The resulting particles showed remarkable activity, inhibiting all ice growth below 1 mg.mL-1. These results are significant as they show that Nature’s approach to hyperactivity, based upon aggregation/self-assembly, can be mimicked using polymer self-assembly.
Our recent work using a photochemical high-throughput discovery platform to identify new macromolecular cryoprotectants has been published in Macro Letters.
In this study we used a liquid handling system to produce 120 unique terpolymers using photopolymerisation with RAFT agents. These terpolymers were screened using a red blood cell freezing assay to identify the best and the worst polymer cryoprotectants, allowing us to explore the chemical space in a short time frame. Testing the hit polymers with a nucleated cell line demonstrated that the high throughput screen was able predict how well the polymers would perform as cryoprotective agents in a more complex assay. This new high throughput approach will allow us to identify new, potent macromolecular cryoprotectants.
Read the paper here:
New work on using X-rays for assessment of ice recrystallisation inhibition is published in Analyst
X-ray diffraction to probe the kinetics of ice-recrystallisation inhibition
We have a large program to discover and understand macromolecular cryoprotectants and new materials to control ice growth. This is typically assessed by end-point assays using averaged data from a small number of ice crystals. Here we make use of the University of Warwick's excellent X-ray scattering platform to show how we can use a 2-D detector on WAXS (wide-angle x-ray scattering) to monitor ice recrsytallisaton in a bulk sample enabling 100's of crystals to be sampled. This method enables easy access to kinetic information which is challenging to obtain from cryo-microscopy alone.
Our recent work into how macromolecular antifreeze agents interact with ice was recently published in ACS Macroletters.
Enhancement of Macromolecular Ice Recrystallisation Inhibition Activity by Exploiting Depletion Forces
We have shown that the ice recrystallisation inhibition of poly(vinyl alcohol) can be improved by the addition of other, inactive macromolecules, in this case poly(ethylene glycol). The additive causes a depletant effect in the liquid channels between ice grains, driving PVA out of solution and on to the the ice crystal surface.
These results give greater insight into the mechanism behind poly(vinyl alcohol)'s antifreeze activity and open up a new suite of tools we can use to make potent antifreeze/cryopreservation agents.
Our perspective article on polyampholytes as emerging macromolecular cryoprotectants has been published in Biomacromolecules.
Polyampholytes as Emerging Macromolecular Cryoprotectants
This perspective is the first review type article on polyampholytes as cryoprotectants and it covers important contributions from both our group and other global leaders in this area.In this perspective we summarise typical methods of cryopreservation for mammalian cells and highlight some of the specific challenges. We then discuss the synthesis and properties of polyampholytes as macromolecular cryoprotectants and detail ways in which polyampholytes have been used to enhance mammalian cell cryopreservation. We further hypothesise about their specific function and how this exciting new field will develop.