Facile and Multiple Replication of Superhydrophilic–Superhydrophobic Patterns Using Adhesive Tape

Surfaces patterned with both hydrophilic and hydrophobic regions are useful in a variety of applications. For example, they can be used as surface tension-confined microchannels, in paper-based microfluidics, or for patterning cells. To create a new patterned substrate, usually the entire experimental procedure must be repeated, which can be time-consuming and laborious. In this paper, we present a simple and fast method that allows the transfer of superhydrophilic–superhydrophobic micropatterns in porous polymer films onto adhesive tape. Replicating patterns using adhesive tape is economical, as the fabrication of one patterned substrate can be used to create multiple copies of the micropatterns, which can then be used for several different experiments. We demonstrate that at least twelve consecutive copies can be made from 125 μm-thick patterned polymer films. Since the polymer film is transferred to adhesive tape, which is flexible, the copies can be used on curved surfaces and they can also be cut into different shapes and sizes. We also demonstrate an application of the replicated patterned polymer surfaces as a substrate for reverse cell transfection experiments

Temperature-Induced Inversion of the Elution Order of Enantiomers in Gas Chromatography: <i>N</i>-Ethoxycarbonyl Propylamides and <i>N</i>-Trifluoroacetyl Ethyl Esters of α-Amino Acids on Chirasil-Val-C₁₁ and Chirasil-Dex Stationary Phases

Inversion of the elution order of enantiomers caused by enthalpy−entropy compensation at the isoenantioselective temperature (Tiso) was experimentally observed by gas chromatography on the diamide-type chiral stationary phase (CSP), Chirasil-l-Val-C11, with N-ethoxycarbonyl propylamide (ECPA) derivatives of a number of α-amino acids. For the first time, a clear visual representation of the increase of the apparent enantioseparation factor αapp from 1.00 to 1.08 as the temperature is raised from 120 to 170 °C is presented. The increase of αapp is accompanied by a concomitant reduction of the retention factors of the enantiomers. The Tiso values were in the range from 110 to 130 °C depending on the nature of the α-amino acid. On the contrary, the Tiso values of the N(O)-trifluoroacetyl ethyl ester derivatives (TFA-Et) of the same α-amino acids were ∼80° higher than that of ECPA derivatives. The comprehensive thermodynamic investigation of the enantioseparation of ECPA and TFA-Et derivatives of valine and alanine using the retention increment method showed that the ΔL,D(ΔH) difference between the diastereomeric selector−selectand associates was almost the same for ECPA and TFA-Et derivatives despite a much stronger bonded selector−selectand association taking place for the ECPA derivatives. On the other hand, the ΔL,D(ΔS) values were found to be more negative in the case of ECPA derivatives, resulting in the unusually low values of Tiso. A temperature-dependent inversion of the elution order of enantiomers was also observed on the cyclodextrin-type CSP, Chirasil-Dex, with TFA-Et derivatives of several α-amino acids. The Tiso values were in the range from 20 to 170 °C depending on the nature of the α-amino acid. The results obtained demonstrate the necessity to conduct temperature-dependent studies in order to optimize the enantiomeric separation of single racemates isothermally or of mixtures of racemates in temperature-programmed runs using enantioselective GC. It is also shown that consideration of the elution order of enantiomers and the value of the apparent enantioseparation factor αapp alone, without performing temperature-dependent measurements, can easily lead to wrong conclusions regarding the enantiorecognition mechanism

Bacterial Cellulose Promotes Long-Term Stemness of mESC

Stem cells possess unique properties, such as the ability to self-renew and the potential to differentiate into an organism’s various cell types. These make them highly valuable in regenerative medicine and tissue engineering. Their properties are precisely regulated in vivo through complex mechanisms that include multiple cues arising from the cell interaction with the surrounding extracellular matrix, neighboring cells, and soluble factors. Although much research effort has focused on developing systems and materials that mimic this complex microenvironment, the controlled regulation of differentiation and maintenance of stemness in vitro remains elusive. In this work, we demonstrate, for the first time, that the nanofibrous bacterial cellulose (BC) membrane derived from Komagataeibacter xylinus can inhibit the differentiation of mouse embryonic stem cells (mESC) under long-term conditions (17 days), improving their mouse embryonic fibroblast (MEF)-free cultivation in comparison to the MEF-supported conventional culture. The maintained cells’ pluripotency was confirmed by the mESCs’ ability to differentiate into the three germ layers (endo-, meso-, and ectoderm) after having been cultured on the BC membrane for 6 days. In addition, the culturing of mESCs on flexible, free-standing BC membranes enables the quick and facile manipulation and transfer of stem cells between culture dishes, both of which significantly facilitate the use of stem cells in routine culture and various applications. To investigate the influence of the structural and topographical properties of the cellulose on stem cell differentiation, we used the cellulose membranes differing in membrane thickness, porosity, and surface roughness. This work identifies bacterial cellulose as a novel convenient and flexible membrane material enabling long-term maintenance of mESCs’ stemness and significantly facilitating the handling and culturing of stem cells

Bacterial Cellulose Promotes Long-Term Stemness of mESC

Stem cells possess unique properties, such as the ability to self-renew and the potential to differentiate into an organism’s various cell types. These make them highly valuable in regenerative medicine and tissue engineering. Their properties are precisely regulated in vivo through complex mechanisms that include multiple cues arising from the cell interaction with the surrounding extracellular matrix, neighboring cells, and soluble factors. Although much research effort has focused on developing systems and materials that mimic this complex microenvironment, the controlled regulation of differentiation and maintenance of stemness in vitro remains elusive. In this work, we demonstrate, for the first time, that the nanofibrous bacterial cellulose (BC) membrane derived from Komagataeibacter xylinus can inhibit the differentiation of mouse embryonic stem cells (mESC) under long-term conditions (17 days), improving their mouse embryonic fibroblast (MEF)-free cultivation in comparison to the MEF-supported conventional culture. The maintained cells’ pluripotency was confirmed by the mESCs’ ability to differentiate into the three germ layers (endo-, meso-, and ectoderm) after having been cultured on the BC membrane for 6 days. In addition, the culturing of mESCs on flexible, free-standing BC membranes enables the quick and facile manipulation and transfer of stem cells between culture dishes, both of which significantly facilitate the use of stem cells in routine culture and various applications. To investigate the influence of the structural and topographical properties of the cellulose on stem cell differentiation, we used the cellulose membranes differing in membrane thickness, porosity, and surface roughness. This work identifies bacterial cellulose as a novel convenient and flexible membrane material enabling long-term maintenance of mESCs’ stemness and significantly facilitating the handling and culturing of stem cells

Combinatorial Synthesis and High-Throughput Screening of Alkyl Amines for Nonviral Gene Delivery

Efficient delivery of plasmid DNA and siRNA into cells is essential for biological and biomedical research. Although significant efforts have been made to develop efficient nonviral vectors, such as cationic lipids and polymers, most of the vectors require multistep synthesis, which complicates both fast structural optimizations and combinatorial synthesis of such vectors. Here, we present a facile, single-step method based on an alkylation of amines, allowing for the fast parallel synthesis of libraries of cationic lipid-like molecules (lipidoids). We exploited the method to synthesize 200 lipidoids, which were screened for their transfection efficiency in HEK293T cells. The screen resulted in about 2% of new lipidoids capable of efficient cell transfection similar or higher than the efficiency of Lipofectamine 2000. In addition, we observed an enhancement of cellular transfection by combining single- with double-chain lipidoids, which was attributed to the different roles of the single- and double-tailed lipids in the mixed liposomes

One-Pot Parallel Synthesis of Lipid Library via Thiolactone Ring Opening and Screening for Gene Delivery

Efficient delivery of nucleic acids into cells is of great interest in the field of cell biology and gene therapy. Despite a lot of research, transfection efficiency and structural diversity of gene-delivery vectors are still limited. A better understanding of the structure–function relationship of gene delivery vectors is also essential for the design of novel and intelligent delivery vectors, efficient in “difficult-to-transfect” cells and in vivo clinical applications. Most of the existing strategies for the synthesis of gene-delivery vectors require multiple steps and lengthy procedures. Here, we demonstrate a facile, three-component <i>one-pot</i> synthesis of a combinatorial library of 288 structurally diverse lipid-like molecules termed “lipidoids” via a thiolactone ring opening reaction. This strategy introduces the possibility to synthesize lipidoids with hydrophobic tails containing both unsaturated bonds and reducible disulfide groups. The whole synthesis and purification are convenient, extremely fast, and can be accomplished within a few hours. Screening of the produced lipidoids using HEK293T cells without addition of helper lipids resulted in identification of highly stable liposomes demonstrating ∼95% transfection efficiency with low toxicity

Single-Tailed Lipidoids Enhance the Transfection Activity of Their Double-Tailed Counterparts

Cationic lipid-like molecules (lipidoids) are widely used for in vitro and in vivo gene delivery. Nearly all lipidoids developed to date employ double-tail or multiple-tail structures for transfection. Single-tail lipidoids are seldom considered for transfection as they have low efficiency in gene delivery. So far, there is no detailed study on the contribution to transfection efficiency of single-tail lipidoids when combined with standard double-tail lipidoids. Here, we use combinatorial chemistry to synthesize 17 double-tail and 17 single-tail lipidoids using thiol–yne and thiol–ene click chemistry, respectively. HEK 293T cells were used to analyze transfection efficiency by fluorescence microscopy and calculated based on the percentage of cells transfected. The size and zeta potential of liposomes and lipoplexes were characterized by dynamic light scattering (DLS). Intracellular DNA delivery and trafficking was further examined using confocal microscopy. Our study shows that combining single with double-tail lipidoids increases uptake of lipoplexes, as well as cellular transfection efficiency

Printable Superhydrophilic–Superhydrophobic Micropatterns Based on Supported Lipid Layers

A new and simple method for creating superhydrophilic micropatterns on a superhydrophobic surface is demonstrated. The method is based on printing an “ink”, an ethanol solution of a phospholipid, onto a porous superhydrophobic surface and, thus, is compatible with a variety of commonly available printing techniques

Bioinspired Strategy for Controlled Polymerization and Photopatterning of Plant Polyphenols

Plant-derived polyphenols have been widely used to design new multifunctional materials for particle and surface modification. The lack of temporal and spatial control over the oxidation and deposition processes, however, limits possible applications of this diverse class of natural compounds. Autoxidation and deposition of phenolic compounds are uncontrollably triggered by basic pH and oxygen. In this project, inspired by the properties of natural antioxidants to scavenge free radicals and reactive oxygen species (ROS), we propose a method to effectively control the autoxidation and deposition of plant phenolic compounds under basic conditions. We demonstrate that natural antioxidants can inhibit autoxidation of plant polyphenols in basic pH in dark environment. However, UV irradiation of these solutions leads to on-demand temporal and spatial polymerization and deposition of polyphenols. This bioinspired method was used to demonstrate the controlled polymerization, and micropatterning on flat surfaces and inside microfluidic channels opening the way to well-defined 2D coatings based on natural polyphenols and introducing a new simple path to the fabrication of bioinspired functional materials, with potential applications in a wide range of fields

Further Dimensions for Sensing in Biofluids: Distinguishing Bioorganic Analytes by the Salt-Induced Adaptation of a Cucurbit[7]uril-Based Chemosensor

Insufficient binding selectivity of chemosensors often renders biorelevant metabolites indistinguishable by the widely used indicator displacement assay. Array-based chemosensing methods are a common workaround but require additional effort for synthesizing a chemosensor library and setting up a sensing array. Moreover, it can be very challenging to tune the inherent binding preference of macrocyclic systems such as cucurbit­[n]­urils (CBn) by synthetic means. Using a novel cucurbit[7]­uril-dye conjugate that undergoes salt-induced adaptation, we now succeeded in distinguishing 14 bioorganic analytes from each other through the facile stepwise addition of salts. The salt-specific concentration-resolved emission provides additional information about the system at a low synthetic effort. We present a data-driven approach to translate the human-visible curve differences into intuitive pairwise difference measures. Ion mobility experiments combined with density functional theory calculations gave further insights into the binding mechanism and uncovered an unprecedented ternary complex geometry for CB7. TThis work introduces the non-selectively binding, salt-adaptive cucurbit­[n]­uril system for sensing applications in biofluids such as urine, saliva, and blood serum