Genetic Network Design Automation with LOICA.

Genetic design automation (GDA) is the use of computer-aided design (CAD) in designing genetic networks. GDA tools are necessary to create more complex synthetic genetic networks in a high-throughput fashion. At the core of these tools is the abstraction of a hierarchy of standardized components. The components' input, output, and interactions must be captured and parametrized from relevant experimental data. Simulations of genetic networks should use those parameters and include the experimental context to be compared with the experimental results.This chapter introduces Logical Operators for Integrated Cell Algorithms (LOICA), a Python package used for designing, modeling, and characterizing genetic networks using a simple object-oriented design abstraction. LOICA represents different biological and experimental components as classes that interact to generate models. These models can be parametrized by direct connection to the Flapjack experimental data management platform to characterize abstracted components with experimental data. The models can be simulated using stochastic simulation algorithms or ordinary differential equations with varying noise levels. The simulated data can be managed and published using Flapjack alongside experimental data for comparison. LOICA genetic network designs can be represented as graphs and plotted as networks for visual inspection and serialized as Python objects or in the Synthetic Biology Open Language (SBOL) format for sharing and use in other designs.

Flapjack: Data Management and Analysis for Genetic Circuit Characterization.

Flapjack presents a valuable solution for addressing challenges in the Design, Build, Test, Learn (DBTL) cycle of engineering synthetic genetic circuits. This platform provides a comprehensive suite of features for managing, analyzing, and visualizing kinetic gene expression data and associated metadata. By utilizing the Flapjack platform, researchers can effectively integrate the test phase with the build and learn phases, facilitating the characterization and optimization of genetic circuits. With its user-friendly interface and compatibility with external software, the Flapjack platform offers a practical tool for advancing synthetic biology research.This chapter provides an overview of the data model employed in Flapjack and its hierarchical structure, which aligns with the typical steps involved in conducting experiments and facilitating intuitive data management for users. Additionally, this chapter offers a detailed description of the user interface, guiding readers through accessing Flapjack, navigating its sections, performing essential tasks such as uploading data and creating plots, and accessing the platform through the pyFlapjack Python package.

Enhancing Fatigue Resistance in Asphalt Mixtures with a Novel Additive Derived from Recycled Polymeric Fibers from End-of-Life Tyres (ELTs).

Waste-tire textile fibers (WTTF) represent a challenge for the recycling industry since there are currently very few alternatives for their use. In this study, an evaluation of the effect of a new additive developed in two granular formats from WTTF on the fatigue behavior of asphalt mixtures was performed. For the first format of the WTTF-based additive, its effect was evaluated on hot-mix asphalt (HMA), while for the second format of the additive, the effects were evaluated on stone mastic asphalt (SMA). This second format represents an alternative that allows for the total replacement of the cellulose stabilizing additive used in the reference mix. The evaluation of fatigue damage in the mixes was performed using the four-point bending beam (4PB) test specified in European standard EN 12697-24. The test results show that the asphalt mixtures manufactured with WTTF-based additives exhibited a higher capacity to resist load cycles before failure compared to the reference mixtures. Likewise, once the asphalt mixtures were evaluated in a pavement structure by means of an empirical mechanistic analysis, the pavement structures composed of asphalt mixtures with WTTF-based additives showed significant improvements in their durability for the different load axes evaluated. For an average thickness of 15 cm of asphalt mix of a pavement-type structure, the use of the WTTF additive increases the durability of the structures by up to 129% and 112% compared to the HMA and SMA reference mixtures, respectively. These results show that both formats of the WTTF-based admixture improve the fatigue damage resistance of the HMA and SMA asphalt mixtures.

Towards Precision Medicine in Clinical Practice: Alinity C vs. UHPLC-MS/MS in Plasma Aripiprazole Determination.

Therapeutic drug monitoring improves the benefit-risk balance of antipsychotic therapy. Ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) is considered the gold-standard method for measuring plasma drug concentrations; however, the Alinity C system has emerged as a promising alternative. This is the first study aimed at comparing UHPLC-MS/MS versus Alinity C in measuring plasma concentrations of aripiprazole and dehydroaripiprazole. A total of 86 plasma samples were analyzed. The active moiety of aripiprazole was measured in 60 samples using both systems and 26 samples were analyzed twice using Alinity C with an intermediate period of 6 months to assess its reproducibility. Spearman's correlation revealed a good association between the two assays (rs = 0.96) and no significance differences were found by McNemar's test when classifying samples between infra-, supra- and therapeutic ranges. Passing-Bablock regression showed a good correlation among methods (rs = 0.93) and a slope of 1.12 indicating a slight tendency of Alinity C to measure higher values than UHPLC-MS/MS. In addition, a good intra-method correlation across the two sequential analyses with Alinity C was obtained (rs = 0.99). Nonetheless, clinical decisions could be different in 15% of the cases depending on the chosen method. No differences were found in active moiety determination by Alinity C depending on the concentration of aripiprazole and dehydroaripiprazole of the samples.

Standardized Representation of Parts and Assembly for Build Planning.

The design and construction of genetic systems, in silico, in vitro, or in vivo, often involve the handling of various pieces of DNA that exist in different forms across an assembly process: as a standalone "part" sequence, as an insert into a carrier vector, as a digested fragment, etc. Communication about these different forms of a part and their relationships is often confusing, however, because of a lack of standardized terms. Here, we present a systematic terminology and an associated set of practices for representing genetic parts at various stages of design, synthesis, and assembly. These practices are intended to represent any of the wide array of approaches based on embedding parts in carrier vectors, such as BioBricks or Type IIS methods (e.g., GoldenGate, MoClo, GoldenBraid, and PhytoBricks), and have been successfully used as a basis for cross-institutional coordination and software tooling in the iGEM Engineering Committee.

Experimental Data Connector (XDC): Integrating the Capture of Experimental Data and Metadata Using Standard Formats and Digital Repositories.

Accelerating the development of synthetic biology applications requires reproducible experimental findings. Different standards and repositories exist to exchange experimental data and metadata. However, the associated software tools often do not support a uniform data capture, encoding, and exchange of information. A connection between digital repositories is required to prevent siloing and loss of information. To this end, we developed the Experimental Data Connector (XDC). It captures experimental data and related metadata by encoding it in standard formats and storing the converted data in digital repositories. Experimental data is then uploaded to Flapjack and the metadata to SynBioHub in a consistent manner linking these repositories. This produces complete connected experimental data sets that are exchangeable. The information is captured using a single template Excel Workbook, which can be integrated into existing experimental workflow automation processes and semiautomated capture of results.

Effect of a New Additive Based on Textile Fibres from End-of-Life Tyres (ELT) on the Mechanical Properties of Stone Mastic Asphalt.

Stone Mastic Asphalts (SMA) are asphalt mixes with discontinuous granulometry and a high content of asphalt binder. In order to prevent draindown of the asphalt binder and ensure good performance, these mixes must be strengthened with cellulose or mineral fibres and/or polymer additives. This study was designed to evaluate the effect of a granular additive based on waste tyre textile fibres (WTTF), developed as a replacement for cellulose commercial additives in SMA mixes. Use of the WTTF-based additive will encourage the development of sustainable mixes by recycling a by-product of end-of-life tyres (ELT), which currently constitute a major environmental problem around the world. To this end, in the present experimental study we evaluated the replacement of cellulose-based commercial fibre with different percentages of WTTF-based additive (0%, 50%, 75%, 100%) in an SMA asphalt mix. The following design and performance properties were evaluated: resistance to cracking, stiffness modulus, sensitivity to moisture, and resistance to permanent deformation. The results indicated that replacing 100% of the cellulose commercial additive in the SMA mix by the WTTF-based additive allowed the mix to meet its design properties and showed good performance in the mechanical properties evaluated, with behaviour similar to that of the reference SMA mix.

Functional Synthetic Biology.

Synthetic biologists have made great progress over the past decade in developing methods for modular assembly of genetic sequences and in engineering biological systems with a wide variety of functions in various contexts and organisms. However, current paradigms in the field entangle sequence and functionality in a manner that makes abstraction difficult, reduces engineering flexibility and impairs predictability and design reuse. Functional Synthetic Biology aims to overcome these impediments by focusing the design of biological systems on function, rather than on sequence. This reorientation will decouple the engineering of biological devices from the specifics of how those devices are put to use, requiring both conceptual and organizational change, as well as supporting software tooling. Realizing this vision of Functional Synthetic Biology will allow more flexibility in how devices are used, more opportunity for reuse of devices and data, improvements in predictability and reductions in technical risk and cost.

Cysteamine Eye Drops in Hyaluronic Acid Packaged in Innovative Single-Dose Systems, Part II: Long-Term Stability and Clinical Ocular Biopermanence.

BACKGROUND: Cystinosis is a rare genetic disorder characterized by the accumulation of cystine crystals in several tissues and organs causing, among others, severe eye symptoms. The high instability of cysteamine eye drops makes it difficult to develop formulations with an acceptable shelf life to be prepared in hospital pharmacy departments. Previously, a new compounded formulation of cysteamine eye drops in hyaluronic acid (HA) packaged in innovative single-dose systems was developed. METHODS: Long-term stability at -20 °C of this formulation was studied considering the content of cysteamine, pH, osmolality, viscosity, and microbiological analysis. The oxygen permeability of single-dose containers was also studied and an ocular biopermanence study was conducted in healthy volunteers measuring lacrimal stability and volume parameters. RESULTS: Data confirm that cysteamine concentration remained above 90% for 120 days, all parameters remaining within the accepted range for ophthalmic formulations. The permeability of the containers was reduced over time, while ocular biopermanence was maintained despite the freezing process and storage time. CONCLUSIONS: 0.55% cysteamine hydrochloride formulation in HA and packaged in single-dose containers preserved at -20 °C is stable for 120 days protected from light, presenting high potential for its translation into clinical practice when commercial presentations are not available.

Ampicillin Stability in a Portable Elastomeric Infusion Pump: A Step Forward in Outpatient Parenteral Antimicrobial Therapy.

Outpatient parenteral antimicrobial therapy (OPAT) with continuous infusion pumps is postulated as a very promising solution to treat complicated infections, such as endocarditis or osteomyelitis, that require patients to stay in hospital during extended periods of time, thus reducing their quality of life and increasing the risk of complications. However, stability studies of drugs in elastomeric devices are scarce, which limits their use in OPAT. Therefore, we evaluated the stability of ampicillin in sodium chloride 0.9% at two different concentrations, 50 and 15 mg/mL, in an elastomeric infusion pump when stored in the refrigerator and subsequently in real-life conditions at two different temperatures, 25 and 32 °C, with and without the use of a cooling device. The 15 mg/mL ampicillin is stable for up to 72 h under refrigeration, allowing subsequent dosing at 25 °C for 24 h with and without a cooling device, but at 32 °C its concentration drops below 90% after 8 h. In contrast, 50 mg/mL ampicillin only remains stable for the first 24 h under refrigeration, and subsequent administration at room temperature is not possible, even with the use of a cooling system. Our data support that 15 mg/mL AMP is suitable for use in OPAT if the volume and rate of infusion are tailored to the dosage needs of antimicrobial treatments.

LOICA: Integrating Models with Data for Genetic Network Design Automation.

Genetic design automation tools are necessary to expand the scale and complexity of possible synthetic genetic networks. These tools are enabled by abstraction of a hierarchy of standardized components and devices. Abstracted elements must be parametrized from data derived from relevant experiments, and these experiments must be related to the part composition of the abstract components. Here we present Logical Operators for Integrated Cell Algorithms (LOICA), a Python package for designing, modeling, and characterizing genetic networks based on a simple object-oriented design abstraction. LOICA uses classes to represent different biological and experimental components, which generate models through their interactions. These models can be parametrized by direct connection to data contained in Flapjack so that abstracted components of designs can characterize themselves. Models can be simulated using continuous or stochastic methods and the data published and managed using Flapjack. LOICA also outputs SBOL3 descriptions and generates graph representations of genetic network designs.

Study of the Effect of the Use of Asphalt Binders Modified with Polymer Fibres from End-of-Life Tyres (ELT) on the Mechanical Properties of Hot Mix Asphalt at Different Operating Temperatures.

Processing of end-of-life tyres (ELT) produces polymer fibres (PFELT) as a by-product. PFELT currently presents a challenge to the recycling industry, due to the increasing numbers of ELT and lack of alternatives for the re-use of this material. The object of this investigation was to propose an alternative for re-using PFELT, in order to improve the performance properties of hot mix asphalt (HMA). This study enabled us to understand the relation between the addition of polymer fibre to the aggregate-binder matrix of the HMA in depth, as well as its effects on the mechanical properties of the resulting asphalt mix. To do this, we first made a physical and chemical characterization of the PFELT (TGA, SEM, polarized light, and fluorescence microscopy), establishing a modification methodology using two asphalt binders (CA-24 and CA-14) and three PFELT contents (0.1%, 0.3%, and 0.5%). The HMA was designed using the Marshall method. The mechanical performance of the HMA was evaluated in a range of operating temperatures, from -10 °C to 50 °C, observing the following properties: (1) resistance to thermal cracking; (2) stiffness modulus; (3) indirect tensile strength; and (4) resistance to permanent deformation. The results show that the addition of 0.3% and 0.5% of PFELT to the asphalt binder significantly improved the mechanical performance properties of the mixes studied, with a greater effect at high operating temperatures; the resistance to permanent deformation increased by more than 30%.

Accurate characterization of dynamic microbial gene expression and growth rate profiles.

Genetic circuits are subject to variability due to cellular and compositional contexts. Cells face changing internal states and environments, the cellular context, to which they sense and respond by changing their gene expression and growth rates. Furthermore, each gene in a genetic circuit operates in a compositional context of genes which may interact with each other and the host cell in complex ways. The context of genetic circuits can, therefore, change gene expression and growth rates, and measuring their dynamics is essential to understanding natural and synthetic regulatory networks that give rise to functional phenotypes. However, reconstruction of microbial gene expression and growth rate profiles from typical noisy measurements of cell populations is difficult due to the effects of noise at low cell densities among other factors. We present here a method for the estimation of dynamic microbial gene expression rates and growth rates from noisy measurement data. Compared to the current state-of-the-art, our method significantly reduced the mean squared error of reconstructions from simulated data of growth and gene expression rates, improving the estimation of timing and magnitude of relevant shapes of profiles. We applied our method to characterize a triple-reporter plasmid library combining multiple transcription units in different compositional and cellular contexts in Escherichia coli. Our analysis reveals cellular and compositional context effects on microbial growth and gene expression rate dynamics and suggests a method for the dynamic ratiometric characterization of constitutive promoters relative to an in vivo reference.

Effect of Biochar from Oat Hulls on the Physical Properties of Asphalt Binder.

The purpose of this study was to verify the feasibility of using biochar from oat hulls (BO) as a potential bio-modifier to improve the physical properties of conventional asphalt binder. The BO and asphalt binder were characterized by confocal (fluorescence) laser microscopy, scanning electron microscopy and Fourier transform infrared spectroscopy. Then, an asphalt binder modification procedure was established and modifications with 2.5, 5.0 and 7.5% of BO on the weight of the asphalt binder were evaluated, using a particle size < 75 µm. The physical properties of the evaluated modified asphalt binder with BO were: rotational viscosity in original and aged state, aging index, Fraass breaking point, softening point, penetration, penetration rate and storage stability. The results indicated that the BO has a porous structure, able to interact with the asphalt binder by C=O and C=C bonds. In addition, modification of the asphalt binder with BO increases the rotational viscosity related to high-temperature rutting resistance. The results obtained from the Fraass breaking point and softening point indicated that the use of BO extends the viscoelastic range of the asphalt binder. In addition, the evaluated modifications present low susceptibility to aging and good storage stability.

Development of a New Additive Based on Textile Fibers of End-of-Life Tires (ELT) for Sustainable Asphalt Mixtures with Improved Mechanical Properties.

End-of-life tires (ELT) are a worldwide problem. Rubber, steel, and different textile fibers are the by-products of ELT. Unlike rubber and steel, waste tire textile fibers (WTTF) are disposed of in landfills or burned. This study developed an additive made with WTTF to be incorporated into conventional hot mix asphalt (HMA), and its performance properties were evaluated. First, a characterization of the WTTF used was made and a manufacture protocol was established. Then, a reference HMA was designed and mixtures with different addition percentages (2%, 5% and 8%) of the WTTF-based additive were evaluated. The mechanical properties studied were stiffness modulus, moisture susceptibility, rutting resistance, stripping, and cracking resistance. The results indicated that the addition of the 2% and 5% WTTF-based additive improved these performance properties. Moreover, all addition percentages of the WTTF-based additive evaluated demonstrated a decrease of over 29% in permanent deformation according to the Hamburg Wheel Tracking Test. Thus, the use of the WTTF would not only be valuing a waste, but an asphalt mixture with improved properties would be obtained, contributing to the circular economy by reusing a material and prolonging the useful life of the asphalt mixture.

Stability of insulin eye drops in the treatment of refractory corneal ulcers.

OBJECTIVE: To determine and compare the physicochemical and microbiological stability of two 25 IU/mL insulin eye drop formulations made with normal saline and a balanced salt solution, respectively, stored for 120 days under various conditions. METHOD: Eye drops were compounded in triplicate with 100 IU/mL Actrapid®  insulin and either normal saline or a balanced salt solution as vehicles, and  they were stored alternatively at room temperature (25 °C), in a refrigerator  (2-8 °C) or in a freezer (-20 °C) for 120 days. Insulin concentrations were  determined by ultra-high resolution liquid chromatography, and osmolality and  pH values were measured at days 0, 3, 7, 15, 30, 60, 90 and 120. Likewise,  samples were extracted for microbiological studies on days 0, 30, 60, 90 and  120. RESULTS: The formulation made with normal saline maintained insulin concentrations above 90% of the baseline level after 120 days across all  temperature conditions. In the case of the balanced salt solution- based eye drops, insulin concentration when stored at room temperature or in the freezer remained stable after 120 days, although insulin concentration when stored in the refrigerator fell below 90% on day 90 of the  study. Osmolality and pH values remained constant in both formulations and across all storage conditions. No microbiological growth was observed in any of the samples. CONCLUSIONS: 25 IU/mL insulin eye drops made with normal saline remain  stable for 120 days whether they are stored at room temperature, in a  refrigerator or in a freezer, provided that they are protected from light. When  made with a balanced salt solution, they remain stable for 120 days at room  temperature and in a freezer, their shelf life being reduced to 90 days in the  case of storage in a refrigerator.

OBJETIVO: Determinar y comparar la estabilidad físico-química y microbiológica de dos colirios de insulina 25 UI/ml elaborados con suero fisiológico o balanced salt solution bajo diferentes condiciones de  conservación durante 120 días.Método: Los colirios se elaboraron por triplicado con insulina Actrapid® 100  Ul/ml y balanced salt solution o suero fisiológico como vehículo, y fueron  conservados a temperatura ambiente (25 °C), en nevera (2-8 °C) o congelador  (­20 °C) durante 120 días. Se determinó la concentración de  insulina mediante cromatografía liquida de ultra alta resolución, la osmolalidad  y el pH a días 0, 3, 7, 15, 30, 60, 90 y 120. Asimismo, se extrajeron muestras  para estudios microbiológicos en los días 0, 15, 30, 60, 90 y 120. RESULTADOS: La formulación elaborada con suero fisiológico mantuvo la  concentración de insulina por encima del 90% con respecto a la inicial tras 120  días de estudio en todas las condiciones de temperatura. En el caso del  colirio elaborado con balanced salt solution, la concentración se mantuvo  estable en ambiente y congelador tras 120 días, aunque en nevera descendió  por debajo del 90% a día 90 de estudio. Los valores de osmolalidad y pH se  mantuvieron constantes en ambas formulaciones y condiciones de  conservación. No se observó crecimiento microbiológico en ninguna de las  muestras retiradas. CONCLUSIONES: El colirio de insulina 25 UI/ml elaborado con suero fisiológico  es estable 120 días, conservado tanto a temperatura ambiente como en nevera  o congelador, protegido de la luz. Con balanced salt solution  permanece estable 120 días a temperatura ambiente y congelador, reduciéndose el periodo de validez a 90 días en el caso de la conservación en nevera.

Cysteamine Eye Drops in Hyaluronic Acid Packaged in Innovative Single-Dose Systems: Stability and Ocular Biopermanence.

Cystinosis is a rare genetic disorder characterized by the accumulation of cystine crystals in different tissues and organs causing, among other symptoms, severe ocular manifestations. Cysteamine eye drops are prepared in hospital pharmacy departments to facilitate access to treatment, for which vehicles that provide adequate biopermanence, as well as adaptable containers that maintain its stability, are required. Difficulties related to cysteamine preparation, as well as its tendency to oxidize to cystamine, show the importance of conducting rigorous galenic characterization studies. This work aims to develop and characterize an ophthalmic compounded formulation of cysteamine prepared with hyaluronic acid and packaged in innovative single-dose systems. For this task, the effect of different storage temperatures and the presence/absence of nitrogen on the physicochemical stability of the formulation and its packaging was studied in a scaled manner, until reaching the optimal storage conditions. The results showed that 0.55% cysteamine, prepared with hyaluronic acid and packaged in single-dose containers, is stable for 30 days when stored at -20 °C. In addition, opening vials every 4 h at room temperature after 30 days of freezing maintains the stability of the cysteamine formulation for up to 16 h. Moreover, ocular biopermanence studies were conducted using molecular imaging, concluding that the biopermanence offered by the vehicle is not affected by the freezing process, where a half-life of 31.11 min for a hyaluronic acid formulation stored for 30 days at -20 °C was obtained, compared with 14.63 min for 0.9% sodium chloride eye drops.

Flapjack: Data Management and Analysis for Genetic Circuit Characterization.

Characterization is fundamental to the design, build, test, learn (DBTL) cycle for engineering synthetic genetic circuits. Components must be described in such a way as to account for their behavior in a range of contexts. Measurements and associated metadata, including part composition, constitute the test phase of the DBTL cycle. These data may consist of measurements of thousands of circuits, measured in hundreds of conditions, in multiple assays potentially performed in different laboratories and using different techniques. In order to inform the learn phase this large volume of data must be filtered, collated, and analyzed. Characterization consists of using this data to parametrize models of component function in different contexts, and combining them to predict behaviors of novel circuits. Tools to store, organize, share, and analyze large volumes of measurement and metadata are therefore essential to linking the test phase to the build and learn phases, closing the loop of the DBTL cycle. Here we present such a system, implemented as a web app with a backend data registry and analysis engine. An interactive frontend provides powerful querying, plotting, and analysis tools, and we provide a REST API and Python package for full integration with external build and learn software. All measurements are associated with circuit part composition via SBOL (Synthetic Biology Open Language). We demonstrate our tool by characterizing a range of genetic components and circuits according to composition and context.

Novel Tunable Spatio-Temporal Patterns From a Simple Genetic Oscillator Circuit.

Multicellularity, the coordinated collective behavior of cell populations, gives rise to the emergence of self-organized phenomena at many different spatio-temporal scales. At the genetic scale, oscillators are ubiquitous in regulation of multicellular systems, including during their development and regeneration. Synthetic biologists have successfully created simple synthetic genetic circuits that produce oscillations in single cells. Studying and engineering synthetic oscillators in a multicellular chassis can therefore give us valuable insights into how simple genetic circuits can encode complex multicellular behaviors at different scales. Here we develop a study of the coupling between the repressilator synthetic genetic ring oscillator and constraints on cell growth in colonies. We show in silico how mechanical constraints generate characteristic patterns of growth rate inhomogeneity in growing cell colonies. Next, we develop a simple one-dimensional model which predicts that coupling the repressilator to this pattern of growth rate via protein dilution generates traveling waves of gene expression. We show that the dynamics of these spatio-temporal patterns are determined by two parameters; the protein degradation and maximum expression rates of the repressors. We derive simple relations between these parameters and the key characteristics of the traveling wave patterns: firstly, wave speed is determined by protein degradation and secondly, wavelength is determined by maximum gene expression rate. Our analytical predictions and numerical results were in close quantitative agreement with detailed individual based simulations of growing cell colonies. Confirming published experimental results we also found that static ring patterns occur when protein stability is high. Our results show that this pattern can be induced simply by growth rate dilution and does not require transition to stationary phase as previously suggested. Our method generalizes easily to other genetic circuit architectures thus providing a framework for multi-scale rational design of spatio-temporal patterns from genetic circuits. We use this method to generate testable predictions for the synthetic biology design-build-test-learn cycle.

Recent Research in Ocular Cystinosis: Drug Delivery Systems, Cysteamine Detection Methods and Future Perspectives.

Cystinosis is a rare genetic disorder characterized by the accumulation of cystine crystals in different tissues and organs. Although renal damage prevails during initial stages, the deposition of cystine crystals in the cornea causes severe ocular manifestations. At present, cysteamine is the only topical effective treatment for ocular cystinosis. The lack of investment by the pharmaceutical industry, together with the limited stability of cysteamine, make it available only as two marketed presentations (Cystaran® and Cystadrops®) and as compounding formulations prepared in pharmacy departments. Even so, new drug delivery systems (DDSs) need to be developed, allowing more comfortable dosage schedules that favor patient adherence. In the last decades, different research groups have focused on the development of hydrogels, nanowafers and contact lenses, allowing a sustained cysteamine release. In parallel, different determination methods and strategies to increase the stability of the formulations have also been developed. This comprehensive review aims to compile all the challenges and advances related to new cysteamine DDSs, analytical determination methods, and possible future therapeutic alternatives for treating cystinosis.