Cross-linked Polyacrylamide Gels Research Articles

Technical note: hydrogel-based mimics of prostate cancer with matched relaxation, diffusion and kurtosis for validating multi-parametric MRI.

Protocol standardization and optimization for clinical translation of emerging quantitative multiparametric (mp)MRI biomarkers of high-risk prostate cancer requires imaging references that mimic realistic tissue value combinations for bias assessment in derived relaxation and diffusion parameters. This work aimed to develop a novel class of hydrogel-based synthetic materials with simultaneously controlled quantitative relaxation, diffusion, and kurtosis parameters that mimic in vivo prostate value combinations in the same spatial compartment and allow stable assemblies of adjacent structures. A set of materials with tunable T2, diffusion, and kurtosis were assembled to create quantitative biomimetic (mp)MRI references. T2 was controlled with variable agarose concentration, monoexponential diffusion by polyvinylpyrrolidone (PVP), and kurtosis by addition of lamellar vesicles. The materials were mechanically stabilized by UV cross-linked polyacrylamide gels (PAG) to allow biomimetic morphologies. The reference T2 were measured on a 3T scanner using multi-echo CPMG, and diffusion kurtosis-with multi-b DWI. Agarose concentration controls T2 values which are nominally independent of PVP or vesicle concentration. For agarose PVP hydrogels, monoexponential diffusion values are a function of PVP concentration and independent of agarose concentration. Compared to free vesicles, for agarose-PAG combined with vesicles, diffusion was predominantly controlled by vesicles and PAG, while kurtosis was affected by agarose and vesicle concentration. Both hydrogel classes achieved image voxel parameter values (T2, Da, Ka) for relaxation (T2: 65-255ms), apparent diffusion (Da: 0.8-1.7μm2/ms), and kurtosis (Ka: 0.5-1.25) within the target literature ranges for normal prostate zones and cancer lesions. Relaxation and diffusion parameters remained stable for over 6months for layered material assemblies. A stable biomimetic mpMR reference based on hydrogels has been developed with a range of multi-compartment diffusion and relaxation parameter combinations observed in cancerous and healthy prostate tissue.

Investigation of diffusion and binding properties of uranium in the diffusive gradients in thin-films technique

Environmental context Monitoring uranium concentrations and speciation in aquatic systems is important for pollution control and for environmental studies. Although an in situ speciation technique based on diffusion and uptake of uranium has been developed, known as DGT, there were uncertainties over some parameters affecting the accuracy of the measurements. This study resolved those uncertainties by investigating diffusion and binding properties of uranium in DGT and provided confidence in monitoring uranium in the environment. Rationale Diffusive gradients in thin-films (DGT) ha been used for uranium speciation measurements since 2006, but interpretational difficulties have arisen due to uncertainties in the pH dependence of U diffusion coefficients (D) within the diffusive gel. This work tested the hypothesis that differences in measured D values between laboratories are due to methodological artifacts. Methodology The properties of uranium binding and diffusion of U within DGT were systematically investigated at different conditions between pH 3–8 and ionic strength from 1 to 500 mmol L−1. Previous uncertainties were attributed to loss of U by adsorption within the diffusion cell, identified by mass balance calculations, and a binding efficiency dependent on the type of binding gel. Results The diffusion coefficient in an agarose cross-linked polyacrylamide gel measured using Metsorb DGT of 5.26 ± 0.17 × 10−6 cm2 s−1 agreed well with both values obtained using a pre-conditioned diffusion cell (< 7% difference) and by modelling. The binding efficiencies for Chelex DGT (BChelex = 0.86–0.89) and Fe-oxide DGT (BFe-oxide = 0.64 were smaller than for Metsorb DGT (BMetsorb = 1). No significant differences were found for BChelex at different pH. Discussion This work demonstrated that the diffusion coefficient of uranium is independent of pH and the binding gel used, provided the binding efficiency (B) is taken into account. These findings indicate that DGT equipped with either Metsorb or Chelex binding gels should be capable of measuring U for a wide range of environmental conditions provided the appropriate diffusion coefficients and binding efficiencies are used.

3D hierarchical porous nitrogen-doped carbon/Ni@NiO nanocomposites self-templated by cross-linked polyacrylamide gel for high performance supercapacitor electrode

Three-dimensional nitrogen-doped carbon network incorporated with nickel@nickel oxide core-shell nanoparticles composite (3D NC/Ni@NiO) has been facilely prepared, self-templated by the cross-linked polyacrylamide aerogel precursor containing NiCl2. Characterizations reveal that the Ni@NiO nanoparticles distribute homogeneously in the 3D nitrogen-doped carbon matrix and the composite is of hierarchical porous structure. When used as supercapacitor electrode in a three-electrode system, the 3D NC/Ni@NiO exhibits enhanced electrical conductivity and excellent electrochemical performance, presenting a high specific capacitance (389F g−1 at 5 mV s−1), good rate capability (276 F g−1 at 100 mV s−1) and outstanding cycling performance (with the capacitance retention of 70.2% after 5000 charge-discharge cycles). This is due to the synergistic effects of conductive metallic nickel, pseudocapacitive nickel oxide as well as in situ nitrogen doping of carbon network. Moreover, an asymmetric supercapacitor (ASC) was fabricated with NC/Ni@NiO as positive electrode and active carbon as negative electrode. The ASC device exhibits a maximum energy density of 19.4 W h kg−1 at a power density of 700 W kg−1 and shows good cycling stability (73.8% capacity retention after 3000 cycles), indicating that it has great promise for practical energy storage and conversion application.

Cross-linked polyacrylamide gel as loss circulation materials for combating lost circulation in high temperature well drilling operation

Cross-linked polymer gel has been broadly used to plug severe lost circulation and to improve the strength of the near-wellbore in natural fracture or carbonate cave strata. Gelation time and gel strength are the key factors for gel plugging. In this study, a cross-linked polymer gel with controlled gelation time and high gel strength was fabricated at high temperature, named HTCMG. It was prepared from an acrylamide monomer, crosslinking agent and initiator by free radical polymerization. To extend gelation time at high temperature, colophony was used here to encapsulate the initiator. This complex was characterized under the Fourier transform infrared spectroscopy (FT-IR), by thermogravimetric analysis (TGA) and under the scanning electronic microscopy (SEM). The effects of pH, temperature, monomer concentration and salt concentration on gelation time were assessed. To examine the viscoelastic property and tensile strength of HTCM, HAKKE Mars 40 and uniaxial compression apparatus were employed, respectively. The plugging property of HTCMG was tested by the core displacement test and high-temperature and high-pressure (HTHP) fracture plugging test. According to the results, HTCMG can form gel in temperature range of 80–150 °C, and the gelation time can be practically adjustable. The results of the core displacement test revealed that the bearing capacity of HTCMG can reach a peak of 9.8 MPa at 150 °C, and it can effectively plug 3 mm-width slot. HTCMG is characterized by a lower plugging time, a higher successful plugging ratio and lower economic cost than normal loss circulation materials, suggesting that HTCM is useful for addressing the lost circulation of high-temperature well, and it is of huge significance to engineering application.

Effect of solvent–matrix interactions on structures and mechanical properties of micelle‐crosslinked gels

ABSTRACTPrevious studies on hydrogels crosslinked by acrylated PEO99–PPO65–PEO99 triblock copolymer (F127DA) micelles demonstrate outstanding strength and toughness, which is attributed to the efficient energy dissipation through the hydrophobic association in the micelles. The current study further focuses on how the solvent property affects the structures and the mechanical properties of F127DA micelle crosslinked polyacrylamide gels. Binary solvents comprised of dimethyl sulfoxide (DMSO) and water are used to adjust the polymer/solvent interactions, which consequently tune the conformations of the polymer chains in the network. The presence of DMSO significantly decreases the strength but increased the stretchability of the gels, whereas the overall tensile toughness remained unchanged. In situ small‐angle X‐ray scattering measurements reveal the deformation of micelles along with the stretching direction. A structure evolution mechanism upon solvent change is proposed, according to the experimental observations, to explain influence of solvent quality on the mechanical properties of the micelle‐crosslinked gels. © 2019 Wiley Periodicals, Inc. J. Polym. Sci., Part B: Polym. Phys. 2019, 57, 473–483

Increase in Local Protein Concentration by Field-Inversion Gel Electrophoresis.

Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and nonspecific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein recovery efficiency. Here, we describe the enhancement of protein separation efficiency for up to twofold in conventional one-dimensional PAG electrophoresis (1D PAGE), two-dimensional (2D) PAGE, and native PAGE by implementing pulses of inverted electric field during gel electrophoresis.

Study on rheology and microstructure of phenolic resin cross-linked nonionic polyacrylamide (NPAM) gel for profile control and water shutoff treatments

Phenolic resin cross-linked polymer gel as a plugging agent for profile control and water shutoff has wide industrial applications. There have been many studies on the gel, but research on the rheology and microstructure has not been systematic. In this study, the rheological properties and microstructure of phenolic resin gel were measured by using a Haake Mars 60 rheometer and a Carl Zeiss Jena cryo-etching electron microscope. Viscosity and viscoelastic moduli of the gel rose constantly and the structure became denser during the gelation process. An increasing polymer or cross-linker concentration increased the viscosity and viscoelastic moduli of the gel. This increase is attributed to a higher polymer concentration contributing to form a stronger membrane structure and a higher cross-linker concentration making the density of the spherical particles higher. The addition of salt and acid affected the stability of the gel. As salinity increased or pH decreased, both the viscosity and viscoelastic moduli of the gel decreased. Combined with the microstructures of the gel systems, this outcome confirmed that salt and acid made the structure of the gel fragile and easily dehydrated. The results showed that phenolic resin gel (gelation temperature: 95 °C; aging time: 8 h) can maintain good stability under a salinity of less than 30,000 mg/L or alkaline conditions with pH > 7. This study can help to understand the gelation mechanism of phenolic resin gel and the impact mechanism of various factors.

Microencapsulation of 2,2′-Azobis(2-methylpropionamide) dihydrochloride initiator using acrylonitrile butadiene styrene as shell for application in lost-circulation control

Gel sealants are often used to recover lost circulation, a problem that plagues drilling engineers throughout many oil and gas development project. Organic cross-linked polymer gel such as cross-linked polyacrylamide gel is one of the most popular and available gel sealant materials. It is of strong sealing ability, but the gelation time is too short to control and ensure safe drilling operation. Initiator used to supply free radical can prolong polyacrylamide gelation time, but the gelation time will dramatically decline with the increase of temperature. Microencapsulation of initiator can increase the gelation time because the concentration of initiator is controlled effectively by microcapsules. This paper proposes microcapsules of an initiator, 2,2′-Azobis(2-methylpropionamide) dihydrochloride (AIBA) encapsulated by Acrylonitrile Butadiene Styrene, prolonging the gelation time of cross-linked polyacrylamide gel. It was prepared via coacervation induced by polydimethylsiloxane (PDMS). The gelation time was prolonged to be eight times greater than that initiated by free AIBA initiator. Fourier transform infrared spectroscopy, thermogravimetric analysis, Ultraviolet-visible spectroscopy, specific surface area analysis, and scanning electron microscopy were used to observe the properties of microcapsules. The results indicated that AIBA was successfully microencapsulated and there were nano-sized pores in the outer surfaces of the microcapsules which serve to control the release of AIBA, and viscosity of PDMS and core/shell ration affected the content of AIBA loaded in the microcapsules. Moreover, it is microencapsulation that makes the concentration of AIBA in water increase stable as a function of time for prolonging the gelation time of sealant system. Computerized tomography (CT) scans revealed that the cross-linked polymer gel system significantly reduces the number of pores inside the cores and enhances pressure-bearing ability. Cross-linked polyacrylamide gel initiated by encapsulated AIBA shows notable potential for recovering lost circulation.

Safety assessment of intra-articular 2.5% polyacrylamide hydrogel for treatment of knee osteoarthritis

Purpose: There is a drought of available effective treatments of knee osteoarthritis (OA). However, new therapeutics and devices are being introduced, such as polyacrylamide gels. This study was conducted to retrospectively assess the safety and patient-reported treatment effect of intra-articular (IA) treatment of knee OA with a proprietary 2.5% cross-linked polyacrylamide gel (PAAG) in patients with knee OA. Methods: The safety evaluation encompassed a sample of patients who had been treated intra-articularly for knee OA symptoms with 2.5 % polyacrylamide hydrogel between March 2010 and February 2017. Safety assessments included the retrospective self-reported adverse events (AEs), medical record reviews, and a clinical examination of the treated knee. If any AEs or discomforts were reported, the patient was asked to describe its duration (“Days”, “Weeks”, “Months”) and severity (“Mild”, “Moderate”, “Severe”). For patients who since treatment had received knee arthroplasty surgery, the surgical records were reviewed for any description of abnormal observations. Patient-reported treatment effect vas measured via a transition questionnaire (Trans-Q), in which the patients scored the current knee status regarding ‘pain’, ‘function’, and ‘global’ in relation to pre-treatment status on a 15 points scale from −7 (‘Much worse’) to 7 (‘Much better’), with 0 being ‘No change’. The assessment was done at average 24 months after first treatment (range 4 to 87 months). Results: Of 126 patients invited to the safety assessment, 91 (72%) accepted the invitation. A majority of the study population (n = 66; 73%) reported no adverse events or discomfort after receiving single or multiple treatments with PAAG. Of the 25 patients who reported discomfort or adverse events, 15 reported a sensation of distention immediately after treatment. Most (n = 14 93%) described this as passing over a few days to weeks. 10 patients (67%) described this as mild. Two patients sought medical assistance (at the study clinic) after treatment for exacerbations of pain and inflammation, which were treated by oral analgesics and arthrocentesis resolving the symptoms within weeks. From the clinical examinations, the investigator found no signs of reduced knee ROM or other abnormalities than what is to be expected in a population of knee OA patients. 15 patients had received knee arthroplasty surgery since treatment of which 14 surgical reports were retrieved. In these we found no unexpected descriptions of abnormalities that could be associated with prior treatment with PAAG. In fact, none of the reports mention the presence of PAAG, unidentified foreign bodies or other findings not expected when inspecting an OA knee. The transition questionnaire describes an overall improvement on pain function and global improvement after treatment with PAAG corresponding to a score of 3 (“somewhat better”) for pain, 2 (“A little better”) for function, and 3 (“somewhat better”) for global. No adverse events causing hospitalizations (serious adverse events), intra-articular infections, or allergic reactions were reported. Conclusions: This retrospective safety assessment found no significant incidence of adverse events or serious adverse events related to the intra-articular treatment with a proprietary 2.5 % cross-linked polyacrylamide gel for the relief of knee osteoarthritis pain and disability. The safety of PAAG must be assessed further in a larger prospective study. However, these initial results suggest that the treatment may be regarded as safe.

Horseradish peroxidase immobilization by copolymerization into cross-linked polyacrylamide gel and its dye degradation and detoxification potential

Owing to the ever-increasing environmental and health impacts associated with the discharge of dye-based effluents, effective remediation of industrial waste have become a top priority for the industrialists and environmental fraternity, around the world. Plant-based peroxidases represent a suitable bio-remediating agent for the effective treatment of original dyes or dye-based colored wastewater effluents. In the present study, horseradish peroxidase was immobilized by copolymerization into cross-linked polyacrylamide gel and investigated its potential for the degradation and detoxification of an azo dye, methyl orange. In the presence of N, N′-methylenebisacrylamide as a cross-linker, polyacrylamide gel at 10% concentration furnished >80% of immobilization yield. The surface morphology of the control (free enzyme) and immobilized enzyme, i.e., horseradish peroxidase cross-linked polyacrylamide gel was characterized using scanning electron microscopy. The high yielded horseradish peroxidase cross-linked polyacrylamide gel concentration, i.e., 10% was used to develop a packed bed reactor and exploited for dye degradation and detoxification purposes. A noteworthy level (>90%) of dye degradation was recorded after polyacrylamide gel cross-linked horseradish peroxidase-catalyzed reaction in a packed bed bioreactor. The biodegradation was further assessed by Fourier-transform infrared spectral analysis. The acute toxicity assays demonstrated that enzyme-based bio-degradation might be used effectively for large-scale environmental remediation of dyes and dyes containing wastewater effluents.

Modeling dynamics of Polyacrylamide Gel in Oil-Water Mixtures: Dissipative Particle Dynamics Approach

ABSTRACTUsing dissipative particle dynamics approach, we model phase separation in a ternary system encompassing cross-linked polyacrylamide (PAM) gel, oil and water. PAM gels are widely used in many applications, from food and cosmetic applications to enhanced oil recovery approaches. We show that the PAM nanogel adsorbs at the oil-water interface and spreads out over this interface for the case of a loosely cross-linked polymer network. Tailoring PAM behavior at the oil-water interfaces by controlling gel’s properties could allow one to alter the properties of oil-water emulsions.

Top-down/Bottom-up Mass Spectrometry Workflow Using Dissolvable Polyacrylamide Gels.

Biologists' preeminent toolbox for separating, analyzing, and visualizing proteins is SDS-PAGE, yet recovering the proteins embedded in these polyacrylamide media as intact species is a long-standing challenge for mass spectrometry. In conventional workflows, protein mixtures from crude biological samples are electrophoretically separated at high-resolution within N,N'-methylene-bis-acrylamide cross-linked polyacrylamide gels to reduce sample complexity and facilitate sensitive characterization. However, low protein recoveries, especially for high molecular weight proteins, often hinder characterization by mass spectrometry. We describe a workflow for top-down/bottom-up mass spectrometric analyses of proteins in polyacrylamide slab gels using dissolvable, bis-acryloylcystamine-cross-linked polyacrylamide, enabling high-resolution protein separations while recovering intact proteins over a broad size range efficiently. The inferior electrophoretic resolution long associated with reducible gels has been overcome, as demonstrated by SDS-PAGE of crude tissue extracts. This workflow elutes intact proteins efficiently, supporting MS and MS/MS from proteins resolved on biologists' preferred separation platform.

Synthesis of hydrogel via click chemistry for DNA electrophoresis

This work introduces a novel sieving gel for DNA electrophoresis using a classical click chemistry reaction, the copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC), to cross-link functional polymer chains. The efficiency of this reaction provides, under mild conditions, hydrogels with near-ideal network connectivity and improved physical properties. Hydrogel formation via click chemistry condensation of functional polymers does not involve the use of toxic monomers and UV initiation. The performance of the new hydrogel in the separation of double stranded DNA fragments was evaluated in the 2200 TapeStation system, an analytical platform, recently introduced by Agilent that combines the advantages of CE in terms of miniaturization and automation with the simplicity of use of slab gel electrophoresis. The click gel enables addition of florescent dyes prior to electrophoresis with considerable improvement of resolution and separation efficiency over conventional cross-linked polyacrylamide gels.

A simple, openable and electroosmotic flow-free PMMA chip for electrophoretic titration of moving reaction boundary

Poor control of electroosmotic flow (EOF) often occurs in chip electrophoretic titration (ET) of moving reaction boundary due to both absence of PMMA channel coating and coexistence of acid-base. Also, chip ET still suffers from low reproducibility, instability, disposability and high cost as well. To address these issues, a reusable ET chip (60mm×60mm×5.0mm) was designed via an upper PMMA sheet, a middle silicon membrane and a lower sheet. In the upper sheet, enlarged channel (500μm×200μm×12mm) was fabricated and linked to the anodic and cathodic wells. The upper sheet, middle membrane and lower sheet could be easily assembled together in 20s as an ET chip of supporting 1.4kg tension force, indicating good mechanical stability. Regeneration and reassembling of chip could be finished in 50s. EOF could be simply suppressed by mass chemical cross-linked polyacrylamide gel or physical agarose gel in the enlarged channel. The EOF-free chip could be well used for: (i) acid-base ET; (ii) EOF monitoring; (iii) pKa determination; (iv) protein content analysis. The EOF-free chip showed the following merits: (i) 98% success rate in contrast to less than 30% in normal channels (80μm×20μm×5.0mm); (ii) more than 30 times reuse, indicating reusability and low cost; (iii) less than 10% of relative standard deviation, suggesting fair stability; and (iv) good recovery and accuracy. The developed chip makes great advancement towards real use of chip in acid- base titration, EOF monitoring, pKa determination and protein content analysis.

Scalable lithography from Natural DNA Patterns via polyacrylamide gel.

A facile strategy for fabricating scalable stamps has been developed using cross-linked polyacrylamide gel (PAMG) that controllably and precisely shrinks and swells with water content. Aligned patterns of natural DNA molecules were prepared by evaporative self-assembly on a PMMA substrate, and were transferred to unsaturated polyester resin (UPR) to form a negative replica. The negative was used to pattern the linear structures onto the surface of water-swollen PAMG, and the pattern sizes on the PAMG stamp were customized by adjusting the water content of the PAMG. As a result, consistent reproduction of DNA patterns could be achieved with feature sizes that can be controlled over the range of 40%–200% of the original pattern dimensions. This methodology is novel and may pave a new avenue for manufacturing stamp-based functional nanostructures in a simple and cost-effective manner on a large scale.

Experimental study and application of gels formed by nonionic polyacrylamide and phenolic resin for in-depth profile control

Abstract A phenolic resin cross-linked nonionic polyacrylamide (NPAM) gel used for in-depth profile control was systematically analyzed. Long gelation time and strong gel strength of the gel system was obtained by adjusting NPAM or cross-linker concentration. An increase in temperature and salt concentration accelerated the gelation process and improved the gel strength. Although shearing brings a negative effect on gel performance, the gel system still had strong gel strength which could make it long-term stability in in-depth formation. Differential scanning calorimetry determined that this gel system should be used for oilfield application at temperatures below 143 °C. A compact three-dimensional network structure was formed in the bulk gel system and in porous media which contributed to long-term gel stability in the formation. Sand-pack flow experiments showed that the NPAM gel had a good plugging capacity and could selectively plug high permeability zones. By retention, adsorption, and bridging across the pore throats, the gel systems effectively reduced the permeability of porous media in high permeability zones and diverted fluid into low permeability zones, and thus improved the formation profile. Furthermore, this gel system for in-depth profile control treatment was successfully used in the Xinjiang oilfield of China which provided a reference for other similar high water cut oilfields for water control worldwide.

Enzymatic protein digestion using a dissolvable polyacrylamide gel and its application to mass spectrometry-based proteomics.

Enzymatic protein digestion in polyacrylamide gel has been used for sample pretreatment in mass spectrometry-based proteomics due to its effectiveness in removing contaminants that interfere with sample ionization. However, the difficulty of recovering the digested peptides from the solid gel matrix has been a drawback of this method. Here we have developed a novel in-gel digestion method to enhance peptide recovery using a dissolvable, bis-acrylylcystamine (BAC)-crosslinked polyacrylamide gel. After enzymatic protein digestion in BAC gel, we completely dissolved the gel by reductive treatment with tris-(2-carboxyethyl) phosphine to release the digested peptides from the gel. Our analysis revealed that the reductive dissolution of the BAC gel enhances the peptide recovery, which has a significantly higher protein identification capability than the conventional method, using an insoluble polyacrylamide gel. In addition, protein samples trapped in dehydrated BAC gel were stable at room temperature and reproducible sample recovery was obtained after storage for one week. These results indicate that the proposed method could be an effective tool for conducting sample pretreatment for mass spectrometry-based protein analysis.

A Novel Autocidal Ovitrap for the Surveillance and Control of Aedes aegypti

We describe an inexpensive autocidal ovitrap for Aedes aegypti that uses cross-linked polyacrylamide (PAM) gel as the oviposition substrate. Aedes aegypti females readily laid eggs on PAM gel that had been hydrated with either hay infusion or water. Aedes aegypti larvae that hatched from their eggs desiccated on the surface of the PAM gel. We tested the effects of gel hydration, texture, and type of attractant on trap performance, and compared the capture rates of standard ovitraps with those of PAM gel ovitraps in the field. The results showed that the number of eggs did not vary over a range of gel hydration levels (40-100%) and that more eggs were recovered from ovitraps containing coarse gel than from those containing homogenized gel. The PAM gel hydrated with hay infusion was more attractive to gravid female mosquitoes than gel hydrated with water. In the field, the number of eggs recovered from autocidal ovitraps with PAM gel was similar to that recovered from standard ovitraps with hay infusion.

Fast and selective determination of total protein in milk powder via titration of moving reaction boundary electrophoresis

In this paper, moving reaction boundary titration (MRBT) was developed for rapid and accurate quantification of total protein in infant milk powder, from the concept of moving reaction boundary (MRB) electrophoresis. In the method, the MRB was formed by the hydroxide ions and the acidic residues of milk proteins immobilized via cross-linked polyacrylamide gel (PAG), an acid-base indicator was used to denote the boundary motion. As a proof of concept, we chose five brands of infant milk powders to study the feasibility of MRBT method. The calibration curve of MRB velocity versus logarithmic total protein content of infant milk powder sample was established based on the visual signal of MRB motion as a function of logarithmic milk protein content. Weak influence of nonprotein nitrogen (NPN) reagents (e.g., melamine and urea) on MRBT method was observed, due to the fact that MRB was formed with hydroxide ions and the acidic residues of captured milk proteins, rather than the alkaline residues or the NPN reagents added. The total protein contents in infant milk powder samples detected via the MRBT method were in good agreement with those achieved by the classic Kjeldahl method. In addition, the developed method had much faster measuring speed compared with the Kjeldahl method.

A visual detection of protein content based on titration of moving reaction boundary electrophoresis

A visual electrophoretic titration method was firstly developed from the concept of moving reaction boundary (MRB) for protein content analysis. In the developed method, when the voltage was applied, the hydroxide ions in the cathodic vessel moved towards the anode, and neutralized the carboxyl groups of protein immobilized via highly cross-linked polyacrylamide gel (PAG), generating a MRB between the alkali and the immobilized protein. The boundary moving velocity (VMRB) was as a function of protein content, and an acid–base indicator was used to denote the boundary displacement. As a proof of concept, standard model proteins and biological samples were chosen for the experiments to study the feasibility of the developed method. The experiments revealed that good linear calibration functions between VMRB and protein content (correlation coefficients R>0.98). The experiments further demonstrated the following merits of developed method: (1) weak influence of non-protein nitrogen additives (e.g., melamine) adulterated in protein samples, (2) good agreement with the classic Kjeldahl method (R=0.9945), (3) fast measuring speed in total protein analysis of large samples from the same source, and (4) low limit of detection (0.02–0.15mgmL−1 for protein content), good precision (R.S.D. of intra-day less than 1.7% and inter-day less than 2.7%), and high recoveries (105–107%).