Polyacrylamide gel electrophoresis (PAGE) is widely used for studying proteins and protein-containing objects. However, it is employed most frequently as a qualitative method rather than a quantitative one. In this paper, we show the feasibility of routine digital image acquisition and mathematical processing of electrophoregrams for protein quantification. Both the well-studied model protein molecules (bovine serum albumin) and more complex real-world protein-based products (casein-containing isolate for sports nutrition), which were subjected to mechanical activation in a planetary ball mill to obtain samples characterized by different protein denaturation degrees, were used as study objects. Protein quantification in the mechanically activated samples was carried out. The degree of destruction of individual protein was shown to be higher compared to that of protein-containing mixture after mechanical treatment for an identical amount of time. The methodological approach used in this study can serve as guidance for other researchers who would like to use electrophoresis for protein quantification both in individual form and in protein mixtures. The findings prove that photographic imaging of gels followed by mathematical data processing can be applied for analyzing the electrophoretic data.

In this paper, a prototype of an automatic lab on PCB for agarose preparation and electrophoresis is developed. The dimensions of the device are 38×34 mm2 and includes a conductivity sensor for detecting the TAE buffer (Tris-Acetate-EDTA buffer), a microheater for mixing, a NTC thermistor for controlling the temperature, a LDR sensor for measuring the transparency of the mixture, and two electrodes for performing the electrophoresis. The agarose preparation functions are governed by a microcontroller. The device requires a PMMA structure to define the wells of the agarose gel, and to release the electrodes from the agarose. The maximum voltage and current that the system requires are 40 V to perform the electrophoresis, and 1 A for activating the microheater. The chosen temperature for mixing is 80ºC, with a mixing time of 10 min. In addition, the curing time is about 30 min. This device is intended to be integrated as a part of a larger lab on PCB system for DNA amplification and detection. However, it can be used to migrate DNA amplified in conventional thermocyclers. Moreover, the device can be modified for preparing larger agarose gels and performing electrophoresis in an automatic manner.

To date, most research on amyloid aggregation has focused on describing the structure of amyloids and the kinetics of their formation, while the conformational stability of fibrils remain insufficiently explored. The aim of this work was to investigate the effect of amino acid substitutions on the stability of apomyoglobin (ApoMb) amyloids. Study of amyloid unfolding of ApoMb and its six mutant variants by urea has been carried out. Changes in the structural features of aggregates during unfolding were recorded by the far UV CD and native electrophoresis. It was shown that during the initial stage of denaturation, amyloid's secondary structure partially unfolds. Then, the fibrils undergo dissociation and form intermediate aggregates weighing approximately 1 MDa, which at the last stage of unfolding decompose into 18 kDa monomeic unfolded molecules. The results of unfolding transitions allow concluding that the stability of studied amyloids relative to the intermediate aggregates and of the latter relative to unfolded monomers is higher for ApoMb variants with substitutions that increase the hydrophobicity of the residues. The results presented provide a new insight into the mechanism of stabilization of protein aggregates and can serve as a base for further investigations of the amyloids stability.

It has been known since the 1930’s that all immunoglobulins carry a weak negative charge in physiological solvents. However, there has been no systematic exploration of this fundamental property. Accurate charge measurements have been made using membrane confined electrophoresis in two solvents (pH 5.0 and pH 7.4) on a panel of twelve mAb IgGs, as well as their F(ab’)₂ and Fc fragments. The following observations were made at pH 5.0: 1) the measured charge differs from the calculated charge by ~40 for the intact IgGs, and by ~20 for the Fcs; 2) the intact IgG charge depends on both Fv and Fc sequences, but does not equal the sum of the F(ab)’₂ and Fc charge; 3) the Fc charge is consistent within a class. In phosphate buffered saline, pH 7.4: 1) the intact IgG charges ranged from 0 to -13; 2) the F(ab’)₂ fragments are nearly neutral for IgG1s and IgG2s, and about -5 for some of the IgG4s; 3) all Fc fragments are weakly anionic, with IgG1 < IgG2 < IgG4; 4) the charge on the intact IgGs does not equal the sum of the F(ab’)₂ and Fc charge. In no case is the calculated charge, based on H⁺ binding, remotely close to the measured charge. The charge on IgGs in physiological solvent is sufficiently small to minimize its contribution to thermodynamic nonideality. Some of the mAbs carried a charge in physiological salt that was outside the range observed for serum-purified human poly IgG. To best match physiological properties, a therapeutic mAb should have a measured charge that falls within the range observed for serum-derived human IgGs.

Background: Deficiency of alpha-1 antitrypsin (AAT1) is a rare disorder that represents a significant health threat and early diagnostic a priority issue. We investigate the usefulness of the serum protein electrophoresis (SPE) as an opportunistic screening tool for AAT1 deficiency. Methods: For 6 months, all SPE carried out for any reasons were evaluated in our center. In those with less than 3% of Alpha-1 globulins, AAT1 concentrations were studied. The SERPINA1 gene was subsequently sequenced in those subjects displaying concentrations below 100 mg/dL. Results: Out of the total, 14 patients (0.3%) were identified with low AAT1 concentrations, 11 of them agreeing to enter the study. Of those, mutations in the SERPINA1 gene were discovered in 10 subjects (91%). Heterozygous mutations were detected in 7 patients; 3 had the c.1096G>A mutation (p.Glu366Lys; Pi*Z), 2 the c.863A>T mutation (p.Glu288Val; Pi*S), one the c.221_223delTCT mutation (p.Phe76del; Pi*Malton) having the last one the c.1066G>A (p.Ala356Thr) mutation not previously described. Finally one patient had the c.863A>T mutation in homozygosis whereas two double heterozygous patients c.863A> T/ c.1096G>A were detected. Conclusions: an altered result in the concentration of AAT1 anticipates a mutation in the SERPINA1 gene in a manner close to 91%. The relationship between a decrease in the Alpha-1 globulin band of the SPE and an alteration in the AAT1 concentration is direct in basal states of health. The SPE is presented as a highly sensitive test for opportunistic screening of AAT1 deficiency.

Bi-clonal gammopathy is distinguished by the presence of two different or distinct monoclonal proteins, due to a proliferation of two distinct monoclonal plasma cell clones. A single monoclonal clone can give rise to two different monoclonal proteins. In this report, we present three cases of biclonal multiple myeloma and one case of Waldenstrom's disease that exhibits two biclonal peaks. Serum protein electrophoresis showed two M bands on serum protein electrophoresis.

Pigeonpea (Cajanus cajan (L.) of Fabaceae family belongs to genus Cajanus usually grown in semi-arid tropics of Asia and Oceania, Africa and America. This crop has been a best source for improving food and soil quality amongst farmers. However, its seed have been always questioned for purity. This problem is managed by using polymorphic SSR markers. In present study, a DNA fingerprints generated by seven SSR markers and hybrid testing is performed on Pigeonpea test samples along with parental lines. The seed samples of pigeonpea were germinated in laboratory and three week old leaves samples were used for DNA isolation by CTAB method. A total of 9 alleles were observed in three test samples using three primers out of seven primers. The screening of the allelic data associated with the three cultivated varieties, revealed markers (CcM0246) displayed unique allelic profiles for one variety. Yet, the genetic fingerprinting data is not well resolved to potentially distinguished two bands of hybrid that are merely of 4–8 bp to confirm hybrid testing of seed. Hybrid testing of may be confirmed including more SSR primers prepared from genomic DNA of pigeonpea.

Contamination of the environment with nano- and microplastic particles exerts a threatened impact on the aquatic ecosystems and sustainable catfish aquaculture. The presence of nanoplastics has been found to have a detrimental impact on both aquatic and terrestrial ecosystems. The present study examines the effect of polystyrene nanoplastics on DNA, erythrocytes, oxidative status and renal of catfish in addition to the potential protective effects of Chlorella vulgaris bioremeditation and selenium to hinder this effect. Six equal groups of fish were used as follows: Group 1: served as a control group and received water free from PS NPs; Group 2: was exposed to PS NPs at a concentration of 5mg/L; Group 3: was exposed to PS NPs (5mg/L) + selenium (1mg/kg diet); Group 4: was exposed to PS NPs (5mg/L) + C. vulgaris (25g/kg diet); Group 5: was supplemented with C. vulgaris (25g/kg diet); and Group 6: was supplemented with selenium (1mg/kg diet). The results indicated that PS NPs induced oxidative stress by significantly elevating MDA activities and slightly reducing antioxidant biomarkers, resulting in DNA damage, increased frequency of micronuclei, erythrocyte alterations, and numerous histopathological alterations in kidney tissue. Selenium and C. vulgaris significantly ameliorated the oxidative/antioxidant status, reduced DNA damage, micronucleus frequency, erythrocyte alterations, and morphology of kidney tissue. Nevertheless, further research is needed to evaluate in detail the mechanism behind the toxicity with nano-microplatics in aquatic system.

Burkholderia andropogonis is a severe plant pathogen that can infect more than 50 plant species. Although its genome has been sequenced and annotated, various mechanisms underlying its physiology and pathogenicity remain unknown. Some of these mechanisms can be uncovered through understanding its proteome. In this study, sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry analysis were used to profile the proteome of B. andropogonis. Exactly 538 proteins, which belong to 190 protein families performing 88 different functions were identified. The resulting STRING network analysis identified 28 proteins having highly connected interactions with adenosine triphosphate (ATP)-binding-cassette and (ABC) transport system. To the knowledge of the authors, this is the first comprehensive proteomic study on B. andropogonis.

Transcriptionally controlled tumor protein (TCTP) is a highly conserved protein performing a large number of cellular functions by binding with various partner proteins. The importance of its roles in many diseases requires an assay method to find regulatory compounds. However, the molecular characteristics of TCTP made it difficult to search for chemicals interacting with it. In this study, a tryptophan-based assay method was designed and Y151W mutant TCTP was constructed to search binding chemicals. Since there is no tryptophan in the native sequence of TCTP, the incorporation of tryptophan in the Y151W mutant was very effective to establish the method. A flavonoid library was employed to the assay with the method. With the native and Y151W mutant TCTPs, three flavonoids such as morin, myricetin and isobavachalcone have been found to interact with TCTP. Combined with native gel electrophoresis, the binding region of isobavachalcone was suggested to be the flexible loop of TCTP. This approach can be easily applicable to find binding compounds of proteins with similar molecular characteristics of TCTP.

The formation of a rigid tough interface based on a nacre-like structure in carbon fiber (CF) composites is a promising way to eliminate low delamination resistance. An effective method of coating CFs is electrophoretic deposition (EPD), which, in the case of dissimilar components like graphene oxide (GO) and polymeric glue, usually requires their chemical bonding/ strong interactions. In this work, we focus on chitin nanocrystals (ChNCs), leading to an excellent mechanical performance of artificial nacre, where favorable interactions and bonding with GO were controlled by degrees of deacetylation (5, 15 and 30 %). We have prepared coatings based on GO/ChNCs adducts with 95/5, 90/10,50/50 and 25/75 ratios using optimized EPD conditions (pH, concentration, voltage and time). The prepared materials were characterized using FTIR, TEM, XPS, SEM, DLS, and XRD. SEM evaluation indicates the formation of a homogeneous interlayer, which possesses a fair ability of chemical bonding with the epoxy matrix. Short beam testing of epoxy matrix composites indicates that coating does not decrease stiffness and its relatively low dependence on composition. Therefore, all coatings are promising for a detailed study of delamination resistance using laminate samples. Moreover, facile EPD from water solution/ suspension has a fair potential for industrial applications.

Polyphosphate (polyP) accumulating organisms (PAOs) are the key agent to perform enhanced biological phosphorus removal (EBPR) activity, and intracellular polyP plays a key role in this process. Potential associations between EBPR performance and the polyP structure have been suggested, but are yet to be extensively investigated, mainly due to the lack of established methods for polyP characterization in the EBPR system. In this study, we explored and demonstrated that single-cell Raman spectroscopy (SCRS) can be employed for characterizing intracellular polyPs of PAOs in complex environmental samples such as EBPR systems. The results, for the first time, revealed distinct distribution patterns of polyP length (as Raman peak position) in PAOs in lab-scale EBPR reactors that were dominated with different PAO types, as well as among different full-scale EBPR systems with varying configurations. Furthermore, SCRS revealed distinctive polyP composition/features among PAO phenotypic sub-groups, which are likely associated with phylogenetic and/or phenotypic diversity in EBPR communities, highlighting the possible resolving power of SCRS at the microdiversity level. To validate the observed polyP length variations via SCRS, we also performed and compared bulk polyP length characteristics in EBPR biomass using conventional polyacrylamide gel electrophoresis (PAGE) and solution ³¹P nuclear magnetic resonance (³¹P-NMR) methods. The results are consistent with the SCRS findings and confirmed the variations in the polyP lengths among different EBPR systems. Compared to conventional methods, SCRS exhibited advantages as compared to conventional methods, including the ability to characterize in situ the intracellular polyPs at subcellular resolution in a label-free and non-destructive way, and the capability to capture subtle and detailed biochemical fingerprints of cells for phenotypic classification. SCRS also has recognized limitations in comparison with ³¹P-NMR and PAGE, such as the inability to quantitatively detect the average polyP chain length and its distribution. The results provided initial evidence for the potential of SCRS-enabled polyP characterization as an alternative and complementary microbial community phenotyping method to facilitate the phenotype-function (performance) relationship deduction in EBPR systems.

A novel electrophoretic technique to improve the sensing capabilities of charged particles in solution is presented. The proposed technique may improve the ability of metasurfaces to sense charged particles in solution by forcing them to preferentially sediment within metasurface regions of greatest sensitivity. Such a technique may be useful in various sensing applications, such as in biological, polymer, or environmental sciences, where low concentration particles in solution are of interest. The electrophoretic technique was simulated and experimentally tested using latex nanoparticles in solution. The results suggest that, using this technique, one may theoretically increase the particle density within the metasurface regions of greatest sensitivity by nearly 1900% in comparison to random sedimentation due to evaporation. Such an increase in particle density within the regions of greatest sensitivity may facilitate more precise material property measurements and enhance identification and detection capabilities of metasurfaces to low concentration particles in solution. It was experimentally verified that the electrophoretic technique enabled the preferential gathering of latex nanoparticles within the most sensitive metasurface regions, resulting in 900% - 1700% enhancements in metasurface sensing capabilities.

The mechanisms by which bacteriophage T4 converts the metabolism of its E. coli host to one dedicated to progeny phage production was the subject of decades of intense research in many labs from the 1950’s through the 1980’s. At this point, a wide range of phages are starting to be used therapeutically and in many other applications and also the range of available phage sequence data is skyrocketing. It is thus important to re-explore the extensive available data about the intricacies of the T4 infection process as summarized here, expand it to looking much more broadly at other genera of phages, and explore phage infections using newly-available modern techniques and a range of appropriate environmental conditions.

Aegilops columnaris Zhuk. is tetraploid grass species (2n=4x=28, UcUcXcXc) closely related to Ae. neglecta and growing in Western Asia and a western part of the Fertile Crescent. Genetic diversity of Ae. columnaris was assessed using C-banding, FISH, nuclear and chloroplast (cp)DNA analyses, and gliadin electrophoresis. Cytogenetically Ae. columnaris was subdivided into two groups, C-I and C-II, showing different karyotype structure, C-banding and FISH patterns. Group C-I was more similar to Ae. neglecta. All types of markers revealed significant heterogeneity of the C-II group, although group C-I was also polymorphic. Two chromosomal groups were consistent with plastogroups identified in a current study based on sequencing of three chloroplast intergenic spacer regions. The similarity of group C-I of Ae. columnaris with Ae. neglecta and their distinctness from C-II indicate that divergence of the C-I group was associated with minor genome modifications. Group C-II could emerge from C-I relatively recently, probably due to introgression from another Aegilops species followed by a reorganization of the parental genomes. Most C-II accessions were collected from the very narrow geographic region, and they might originate from a common ancestor. We suggest that the C-II group is at the initial stage of species divergence and undergoing an extensive speciation process.

The extensive use of engineered nanoparticles (ENPs) such as zinc oxide nanoparticles (ZnO NPs) in various commercial fields has spurred significant concern about its possible toxicological effects of ZnO NPs on human health and eco-system. Therefore, it is substantial to understand the aggregation phenomena which could attenuate this effect. This data article provides information on aggregation of ZnO NPs under the influence of various parameters of water chemistry. Moreover, Taguchi orthogonal array L27(313) design matrix was used to evaluate the effect of multiple parameters in aqueous solution. The data were obtained from single beam UV-Vis spectrophotometer (Optizen 2120 UV, Mecasys, Korea) using Optizenview 4.2.5 PC interference software, Fourier transform infrared spectrometer (FT/IR-4700, spectroscopy, JASCO Analytical Instruments, Easton, USA) and particle electrophoresis (NanoZS, Zetasizer, Malvern Instruments Ltd, Worcestershire, UK). The dataset draws attention on dominant parameters influencing aggregation of ZnO NPs in water. The analysis of variance (ANOVA) data revealed that electrolyte concentration, type and natural organic matter (NOM) concentration were the most significant parameters. Furthermore, FT-IR data presents a possible mechanism of ZnO NPs stabilization in the presence of different NOM. This data will be helpful for the development of environmental risk assessment strategies and prediction of fate and mobility of other ENPs in the aquatic environment.

Dysregulation of cartilage homeostasis and the changes in the density and the architecture of the subchondral bone were postulated as a potent mechanically pathological activity contributing to osteoarthritis (OA) pathogenesis. Extracorporeal shockwave therapy (ESWT) is a new, none invasive and effective method in the treatment of animal OA model. In the current study, we demonstrated that shockwave induced the expression of protein-disulfide isomerase-associated 3 (Pdia-3) which is a multifunctional protein hypothesized to be a significant mediator for 1α,25-Dihydroxyvitamin D3 (1α,25(OH)₂D₃) signaling pathway using two-dimensional electrophoresis. Histological analysis and quantitative polymerase chain reaction (qPCR) were verified and observed that the expression of Pdia-3 at 2 weeks was significantly higher than that of any other group at 4 weeks, 8 weeks, and 12 weeks post-shockwave treatment in early OA knee of rat. The other factors of the 1α,25(OH)₂D₃ rapid membrane signaling pathway including extracellular signal-regulated protein kinases 1 (ERK1), osteopontin (OPG), alkaline phosphatase (ALP), and matrix metallopeptidase 13 (MMP13) were measured and significantly increased by qPCR at 2 weeks post-shockwave treatment in early OA knee. Our proteomic data revealed significant Pdia-3 expression in microenvironments of joint tissue that could be actively responded to ESWT, which may potentially regulate biological function of chondrocytes and osteoblasts in the treatment of OA knee.