Slab Gel Research Articles

Association of Serum Proteins Electrophoretic Pattern and Serum Hormones in Women with Spontaneous Pregnancy Loss.

Pregnancy is the state of carrying a developing embryo or fetus within a female body. Once pregnancy is established, a range of endocrinological events appear in its maintenance, finally helping in the successful pregnancy. The complications which are usually observed in pregnancy are gestational diabetes, preeclampsia, preterm labor, and spontaneous pregnancy loss or miscarriage, while 10%-15% of clinically recognized pregnancies terminate into spontaneous miscarriage. Thus, many attempts have been made by different researchers for the diagnosis of high-risk pregnancy on altered protein pattern using placental villous tissue or follicular fluid, but these are difficult to obtain and results of different studies are not constant. This study was designed to identify the association (if any) among serum protein(s) electrophoretic pattern and different serum hormones in normal pregnant women (controls) and gestational age-matched women with spontaneous pregnancy loss (cases). This study was carried out for 1½ year from October 2018 to March 2020 and included 120 participants (60 normal pregnant women and 60 women with spontaneous pregnancy loss) between 20 and 45 years of age with no mean age difference. The electrophoresis of serum was carried out using slab gel electrophoretic unit and serum thyroid-stimulating hormone (TSH), total tri-iodothyronine (TT3), total thyroxine (TT4), prolactin, and beta human chorionic gonadotropin (β-hCG) levels were analyzed using TSOSH AIA analyzer at Adesh University, Bathinda. Significant variations in the expression of proteins with molecular weight around ~150 kDa, ~50 kDa, and ~25 kDa were observed in normal pregnant women and women with spontaneous pregnancy loss. However, the protein band of ~50 kDa was found to be highly expressed in the serum of 1st and 2nd trimester women experiencing spontaneous pregnancy loss. Therefore, selected protein band of ~50 kDa was further processed by ECI-mass spectrophotometry QUAD time of flight and 365 different proteins were found, out of these; 34 proteins were found to be unidentified protein products (Verified using NCBI data base). Further, TT3, total proteins, β-hCG, and prolactin level were found to be low, whereas, TSH was found to be high in women experiencing spontaneous pregnancy loss. However, difference in the level of β-hCG in the 1st trimester and TT4 among normal pregnant women and women with spontaneous pregnancy loss was observed to be statistically insignificant. This study indicated that the evaluation of serum protein variations along with hormonal profile may provide valuable information about high-risk pregnancy. Moreover, the differential expression of proteins in women with spontaneous pregnancy loss can be further explored to develop potential biomarker for the early identification of high-risk pregnancy and appropriate preventive measure.

Real-time and quantitative protein detection via polyacrylamide gel electrophoresis and online intrinsic fluorescence imaging

The detection of intrinsic protein fluorescence is a powerful tool for studying proteins in their native state. Thanks to its label-free and stain-free feature, intrinsic fluorescence detection has been introduced to polyacrylamide gel electrophoresis (PAGE), a fundamental and ubiquitous protein analysis technique, to avoid the tedious detection process. However, the reported methods of intrinsic fluorescence detection were incompatible with online PAGE detection or standard slab gel. Here, we fulfilled online intrinsic fluorescence imaging (IFI) of the standard slab gel to develop a PAGE-IFI method for real-time and quantitative protein detection. To do so, we comprehensively investigated the arrangement of the deep-UV light source to obtain a large imaging area compatible with the standard slab gel, and then designed a semi-open gel electrophoresis apparatus (GEA) to scaffold the gel for the online UV irradiation and IFI with low background noise. Thus, we achieved real-time monitoring of the protein migration, which enabled us to determine the optimal endpoint of PAGE run to improve the sensitivity of IFI. Moreover, online IFI circumvented the broadening of protein bands to enhance the separation resolution. Because of the low background noise and the optimized endpoint, we showcased the quantitative detection of bovine serum albumin (BSA) with a limit of detection (LOD) of 20 ng. The standard slab gel provided a high sample loading volume that allowed us to attain a wide linear range of 0.03–10 μg. These results indicate that the PAGE-IFI method can be a promising alternative to conventional PAGE and can be widely used in molecular biology labs.

Continuous flow delivery system for the perfusion of scaffold-based 3D cultures.

The paper-based culture platform developed by Whitesides readily incorporates tissue-like structures into laboratories with established workflows that rely on monolayer cultures. Cell-laden hydrogels are deposited in these porous scaffolds with micropipettes; these scaffolds support the thin gel slabs, allowing them to be evaluated individually or stacked into thick constructs. The paper-based culture platform has inspired many basic and translational studies, each exploring how readily accessible materials can generate complex structures that mimic aspects of tissues in vivo. Many of these examples have relied on static culture conditions, which result in diffusion-limited environments and cells experiencing pericellular hypoxia. Perfusion-based systems can alleviate pericellular hypoxia and other cell stresses by continually exposing the cells to fresh medium. These perfusion systems are common in microfluidic and organ-on-chip devices supporting cells as monolayer cultures or as 3D constructs. Here, we introduce a continuous flow delivery system, which uses parts readily produced with 3D printing to provide a self-contained culture platform in which cells in paper or other scaffolds are exposed to fresh (flowing) medium. We demonstrate the utility of this device with examples of cells maintained in single cell-laden scaffolds, stacks of cell-laden scaffolds, and scaffolds that contain monolayers of endothelial cells. These demonstrations highlight some possible experimental questions that can be enabled with readily accessible culture materials and a perfusion-based device that can be readily fabricated.

Supported gel slab scaffolds as a three-dimensional cell-based assay platform.

Cell-based assays are heavily relied on in the drug discovery pipeline, quickly pairing down large compound libraries to a manageable number of drug candidates for further characterization and evaluation. Monolayer cultures in which cells are deposited onto the bottom of well plates are the workhorse of many of these screens despite continued evidence of their inability to predict in vivo responses. Three-dimensional (3D) culture platforms can generate tissue-like environments with more representative cellular phenotypes than monolayers but have proven challenging to incorporate into already-developed workflows. Scaffold-based approaches are a tractable means of generating tissue-like environments, supporting cell-laden gels whose preparation is analogous to depositing cells in a well plate. Here, we describe supported gel slab (SGS) scaffolds prepared from commercially available materials, an adhesive spray, and a laser cutter. These cell-containing scaffolds can readily fit into well plates, providing a format compatible with current liquid handlers and analytical instrumentation. The scaffolds enable the evaluation of cellular responses in individual or stacked structures, which contain extracellular matrix-rich microenvironments. With a series of demonstrations, we highlight the utility of the readily assembled SGS scaffolds to quantify cellular responses. These readouts include confocal microscopy, quantifying cellular invasion in Transwell-like and stacked formats, generating multilayered spheroid-on-demand structures capable of providing spatially resolved maps of drug responses, and identifying potential chemotherapies in a screening application.

A model for the contraction kinetics of cytoskeletal gel slabs

Cytoskeletal gels are engineered prototypes that mimic the contractile behavior of a cell in vitro. They are composed of an active polymer matrix and a liquid solvent. Their contraction kinetics is governed by two dynamic phenomena: mechanotransduction (molecular motor activation) and solvent diffusion. In this paper, we solve the transient problem for the simple case of a thin gel slab in uniaxial contraction under two extreme conditions: motor-limited or slow motor (SM) activation regime, and diffusion-limited or fast motor (FM) activation regime. The former occurs when diffusion is much faster than mechanotransduction, while the latter occurs in the opposite case. We observe that in the SM regime, the contraction time scales as t/t0∼(λ/λ0)−3, with t0 being the nominal contraction time, and λ and λ0 being the final and initial stretches of the slab. t0 is proportional to 1/w˙, where w˙ is the average mechanical power generated by the molecular motors per unit reference (dry polymer) volume. In the FM regime, the contraction time scales as t/t1∼(1−λ/λ0)2, with t1 being the nominal contraction time, here proportional to the ratio L2/D, where L is the reference (dry polymer) thickness, and D is the diffusivity of the solvent in the gel. The transition between the SM and FM regimes is defined by a characteristic power density w˙∗, where w˙≪w˙∗ gives the SM regime and w˙≫w˙∗ gives the FM regime. Intuitively, w˙∗ is proportional to D/L2, where, at a given power density w˙, a thinner gel slab (smaller L) or including smaller solvent molecules (higher D) is more likely to be in the SM regime given that solvent diffusion will occur faster than motor activation.

Glowing gels and pipettes aplenty: how do commercial stock image banks portray genetic tests?

News stories and patient-facing material about genetic tests are often illustrated by images, but the content of such images and the messages they propagate are rarely scrutinised. Stock image banks were searched to identify a hundred images relating to genetic tests and analysed using a multimodal critical discourse approach, aiming to identify what the images featured, how they were composed, and what they communicated about genetic testing. We found that images tended to focus on technical aspects of sample processing (for example, pipetting) and drew on older technologies (for example slab gel electrophoresis) when representing data arising from genetic tests. Composition choices like focussing images around pipette tips, or emphasising colour or brightness of electrophoretic bands, represented genetic testing as precise, unambiguous and illuminating. Only 7% of images featured a person having a genetic test, and only one image alluded to communication of genetic results. Current popular visual representations of genetic testing rarely highlight the possibility of uncertain or non-diagnostic outcomes, and may contribute to high public expectations of informativeness and certainty from such tests.

Quadruple UV LED Array for Facile, Portable, and Online Intrinsic Fluorescent Imaging of Protein in a Whole Gel Electrophoresis Chip.

Intrinsic fluorescence imaging (IFI) has been used for the stain-free detection of proteins in slab gel. However, complicated detection setups and small irradiation area limited the development of facile, online, and portable imaging of the whole slab gel. We here designed a quadruple UV LED array to produce even and powerful area light for direct irradiation of gel electrophoresis chip (GEC) at 275 nm. In addition, we only used a filter of 365 nm, a UV camera lens, and a CCD for IFI detection. We integrated the simple detection setup with the small GEC to construct the IFI-GEC device with a portable size of 15 × 15 × 38 cm. We detected three model proteins to demonstrate the good evenness of the LED array and the online imaging of the whole GEC. Furthermore, the reproducible IFI-GEC detection was completed within 10 min and the LOD was as low as 40 ng for lysozyme detection. All results indicated the potential of the IFI-GEC device for online and portable detection of proteins without staining.

Comparative 3-Sample 2D-DIGE Analysis of Skeletal Muscles.

The skeletal muscle proteome consists of a large number of diverse protein species with a broad and dynamic concentration range. Since mature skeletal muscles are characterized by a distinctive combination of contractile cells with differing physiological and biochemical properties, it is essential to determine specific differences in the protein composition of fast, slow, and hybrid fibers. Fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) is a powerful comparative tool to analyze fiber type-specific differences between predominantly fast contracting versus slower twitching muscles. In this chapter, the application of the 2D-DIGE method for the comparative analysis of different subtypes of skeletal muscles is outlined in detail. A standardized proteomic workflow is described, involving sample preparation, protein extraction, differential fluorescence labeling using a 3-CyDye system, first-dimension isoelectric focusing, second-dimension slab gel electrophoresis, 2D-DIGE image analysis, protein digestion, and mass spectrometry.

Assessment of Collagen III as a Non-Invasive Biomarker in HCV Induced Liver Fibrotic Patients

Abstract Introduction/Objective Liver biopsy is still the gold typical for assessing hepatic fibrosis. Non-invasive serum parameters measurement leads to essential changes in diagnosis. This procedure is painful, carries a risk of bleeding and infection, and is accompanied by a high sampling error rate. New biomarkers that can accurately predict the stage of fibrosis are urgently needed. Objectives This study aimed to identify, characterize, and detect serum collagen III in patients with HCV-induced liver fibrosis and healthy subjects. Methods/Case Report The study subjects were divided into two groups; group1: included 118 patients with HCV-induced liver fibrosis (60 males and 58 females). Group2: included 50 healthy comparable control subjects (27 males and 23 females). Collagen III was identified in serum samples of group1 at 70 kDa using collagen III polyclonal antibody and western blot analysis techniques. Additionally, it was purified from serum samples using the electroelution technique from preparative slab gels. The purified antigen gives a single sharp band at 70 kDa when resolved by SDS–PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and stained with coomassie blue. The 70 kDa purified collagen III showed high reactivity towards the collagen III polyclonal antibody. We used the ELISA method to quantify the collagen III, dose-response curve for purified collagen III as a function of the concentration in serum samples of liver fibrosis patients was done. Results (if a Case Study enter NA) The collagen III was identified in 62% of patients, and 38 % were negative collagen III (false negative) in group one. For group two, 96% of total healthy volunteers were negative for collagen III, and 4% were positive for collagen III (false positive). The mean ± SD of serum collagen III concentration was (8.38±3.56) and (3.2±0.59) for groups 1 and 2, respectively, with (P<0.0001). Conclusion The non-invasive assessment of liver fibrosis by collagen III has shown promising diagnostic and prognostic potential.

Mesowestern Blot: Simultaneous Analysis of Hundreds of Submicroliter Lysates.

Western blotting is a widely used technique for molecular-weight-resolved analysis of proteins and their posttranslational modifications, but high-throughput implementations of the standard slab gel arrangement are scarce. The previously developed Microwestern requires a piezoelectric pipetting instrument, which is not available for many labs. Here, we report the Mesowestern blot, which uses a 3D-printable gel casting mold to enable high-throughput Western blotting without piezoelectric pipetting and is compatible with the standard sample preparation and small (∼1 μL) sample sizes. The main tradeoffs are reduced molecular weight resolution and higher sample-to-sample CV, making it suitable for qualitative screening applications. The casted polyacrylamide gel contains 336, ∼0.5 μL micropipette-loadable sample wells arranged within a standard microplate footprint. Polyacrylamide % can be altered to change molecular weight resolution profiles. Proof-of-concept experiments using both infrared-fluorescent molecular weight protein ladder and cell lysate (RIPA buffer) demonstrate that the protein loaded in Mesowestern gels is amenable to the standard Western blotting steps. The main difference between Mesowestern and traditional Western is that semidry horizontal instead of immersed vertical gel electrophoresis is used. The linear range of detection is at least 32-fold, and at least ∼500 attomols of β-actin can be detected (∼29 ng of total protein from mammalian cell lysates: ∼100–300 cells). Because the gel mold is 3D-printable, users with access to additive manufacturing cores have significant design freedom for custom layouts. We expect that the technique could be easily adopted by any typical cell and molecular biology laboratory already performing Western blots.

Electrotransfer of Immunoprobes through Thin-Layer Polyacrylamide Gels.

Hydrogels are important structural and operative components of microfluidic systems, finding diverse utility in biological sample preparation and interrogation. One inherent challenge for integrating hydrogels into microfluidic tools is thermodynamic molecular partitioning, which reduces the in-gel concentration of molecular solutes (e.g., biomolecular regents), as compared to the solute concentration in an applied solution. Consequently, biomolecular reagent access to in-gel scaffolded biological samples (e.g., encapsulated cells, microbial cultures, target analytes) is adversely impacted in hydrogels. Further, biomolecular reagents are typically introduced to the hydrogel via diffusion. This passive process requires long incubation periods compared to active biomolecular delivery techniques. Electrotransfer is an active technique used in Western blots and other gel-based immunoassays that overcomes limitations of size exclusion (increasing the total probe mass delivered into gel) and expedites probe delivery, even in millimeter-thick slab gels. While compatible with conventional slab gels, electrotransfer has not been adapted to thin gels (50-250 μm thick), which are of great interest as components of open microfluidic devices (vs enclosed microchannel-based devices). Mechanically delicate, thin gels are often mounted on rigid support substrates (glass, plastic) that are electrically insulating. Consequently, to adapt electrotransfer to thin-gel devices, we replace rigid insulating support substrates with novel, mechanically robust, yet electrically conductive nanoporous membranes. We describe grafting nanoporous membranes to thin-polyacrylamide-gel layers via silanization, characterize the electrical conductivity of silane-treated nanoporous membranes, and report the dependence of in-gel immunoprobe concentration on transfer duration for passive diffusion and active electrotransfer. Alternative microdevice component layers─including the mechanically robust, electrically conductive nanoporous membranes reported here─provide new functionality for integration into an increasing array of open microfluidic systems.

A modular and reconfigurable open-channel gated device for the electrokinetic extraction of cell-free DNA assays

The high-efficiency separation and extraction of short fragments of cell-free DNA (cfDNA) remain challenging due to their low abundance and short lengths. This study presents a method for separating short cfDNA fragments, with lengths ranging from about 100 to 200 base pairs, from liquid human plasma samples into separable and extractable bands as solid agarose gel slabs. To achieve this, a novel millimeter-scale fluidic device is used for sample handling, transient isotachophoresis, and extraction. The device features open-to-atmosphere liquid chambers that define and manually actuated (i.e., movable) agarose-made gate valve structures. The agarose gates then define discrete zones for buffers, sample injection, DNA pre-concentration via isotachophoresis, size-based gel separation, and DNA-band extraction. As a demonstration of its efficacy, the device is applied to the enrichment and purification of M. tuberculosis genomic DNA fragments spiked in human plasma samples. This purified cfDNA is analyzed using the quantitative polymerase chain reaction (qPCR) of the IS6110 repetitive sequence in the M. tuberculosis genome. The data from this study demonstrates that high sensitivity can be achieved in cfDNA detection, as shown by the comparison with a typical solid-phase extraction method and buffer spiked with cfDNA. Evidence is presented that suggests plasma peptides generated by treatment of the sample with proteinase K acts as endogenous spacer molecules, which improve the resolution and purification of DNA relative to the marker dye and other contaminants that decrease the signal level in qPCR.

Determination of Plasmid Profile Multiple Antibiotics Resistant Pseudomonas aeruginosa Isolated from Clinical Specimens in Healthcare Facilities across Anambra and Imo States Nigeria

The emergence and dissemination of antibiotic resistant plasmids of P. aeruginosa is posing a major public threat and huge concern in hospital facilities. This study was done across Anambra and Imo States with a total number of 100 P. aeruginosa isolates, 50 from each State to determine the plasmid profile of multidrug resistant isolates. Methods of re-identifying P. aeruginosa were based upon cultural methods coupled with biochemical tests. To study the susceptibility of these isolates using disk diffusion method, seven (7) antibiotics were used namely: Piperacillin–tazobactam (100/10 µg), Ceftazidime (30 µg), Amikacin (30 µg), Ciprofloxacin (5 µg), Cefuroxime (30 µg), Ceftriazone (30 µg) and Gentamicin (10 µg). Plasmid extraction was done using alkaline lysis method after growing isolates resistant to more than four antibiotics on a nutrient broth. Agarose gel electrophoresis of plasmid DNA was carried out on 2% agarose gel slab in 1X TAE buffer. The results show that, the nutrient agar at 37˚C aerobically, P. aeruginosa isolates were recovered, which produced greenish-yellow pigment colonies, oxidase was positive and negative for gram stain. In Anambra, the result showed 100% resistance to Cefuroxime, Ceftazidime and Ciprofloxacin, 90% and 86% for Ceftriaxone and Piperacillin-tazobactem, 48% and 40% to Gentamicin and Amikacin respectively. Whereas in Imo state, the result showed 100% resistance to Ceftazidime and Ciprofloxacin, 80% to Ceftriaxone, Cefuroxime and Piperacillin-tazobactem while the least resistance was seen in Amikacin, and Gentamicin. Plasmid size ranging from 100bp to >1000bp was detected from most of the multidrug resistant isolates. Not all the isolates with multidrug resistance were found to possess plasmids. It can be seen from this study that multidrug resistance in P. aeruginosa is not strictly plasmid-dependent (mediated).

A platform method for charge heterogeneity characterization of fusion proteins by icIEF

The charge heterogeneity of fusion proteins can vary dramatically compared with more traditional biopharmaceuticals like monoclonal antibodies, making the characterization of fusion proteins a challenge. A single platform method suitable for the analysis of multiple fusion proteins would reduce method development and streamline production workflows. Here, we develop a platform method to characterize the charge heterogeneity of a variety of fusion protein therapeutics using imaged capillary isoelectric focusing (icIEF). We describe the development of the platform method, and analyze 9 fusion protein therapeutics. The results are reproducible in peak group area percentage and apparent pI determination. We compare the platform icIEF method to traditional slab gel IEF, which is still used in many laboratories for the analysis of fusion proteins. The peak patterns obtained from the icIEF method is comparable to the band patterns of the gel IEF. The platform method can also be used as the starting point if further optimization is needed even when high resolution is required. The platform method described in this study can be applied as an identity and purity assay for fusion proteins in the biopharmaceutical industry.

Online electrophoretic nanoanalysis using miniaturized gel electrophoresis and thermal lens microscopy detection

Online thermal lens microscopy (TLM) coupled with gel electrophoresis (GE) can represent a powerful tool for separating and detecting a wide range of biomaterials. Unlike slab gel electrophoresis (SGE), the proposed method does not require prolonged procedure between separation and detection. In this work, we developed an online monitoring GE system to separate and detect nanosized materials. The design is based on a homemade and cost-effective miniaturized GE chip (MGEC) integrated with real-time TLM detection through microcontroller-based digitization board platform. To validate the feasibility and practicability of the proposed approach, we evaluated its separation capability via employing synthesized Fe3O4-Au core-shell nanoparticles (NPs) which served remarkably for the proof-of-concept. The optimum conditions for the separation process were achieved through optimization of the excitation power as 30 mW, detection position at 24 mm, the concentration of agarose gel 0.5 % w/v, and 37.5 V/cm as the effective electric field strength. The findings showed that two populations of Fe3O4–Au, core-shell, and uncapped Fe3O4 NPs, were effectively separated in less than eleven minutes, demonstrating rapid assessment of the nanomaterial production quality. Moreover, other characterization techniques such as HRTEM and EDX were employed to confirm the presence of the two dissimilar kinds of NPs separated using MGEC-TLM. The sensitivity of the method was demonstrated by determining the limit of detection (23 pM) for 10 nm AuNPs. It is envisaged that our presented system enables rapid, economical, low volume of reagents consumption and high potential analysis for quality test in various bioanalytical and nanotechnological applications.

Robust and easy-to-use microchip electrophoresis within sub-millimeter channels for fast and highly efficient separation

Microchip capillary electrophoresis (MCE) is a powerful technique for rapid separation; however, its acceptance in routine laboratories is still limited. Compromises caused by the efforts for solving different problems, such as reducing its cost of fabrication and ensuring high separation efficiency, undermine the competitiveness of this technology compared to other separation techniques. Contrary to the conventional pursuit of narrow microchannels, this study investigated the suitability of microchips with channels at the sub-millimeter level, targeting the simplification of the overall operation, cost reduction, and robustness improvement. To this effect, we considered the influence of pressurized flow and Joule heating on the separation. The suppression of pressurized flow with viscous solutions was confirmed through a combination of simulations and experimental results, indicating that the buffer viscosity was enough for successful separation. We fabricated channels of 200 μm × 230 μm using computer numerical controlled (CNC) machining and obtained theoretical plate numbers of 4.8 × 105 m−1 and 5.3 × 105 m−1 for fluorescein isothiocyanate (FITC) labeled small molecules and DNA fragments, respectively, with a buffer viscosity of 168 mPa s (0.5 % hydroxypropyl methylcellulose, HPMC). These values are comparable with that of narrow-bore microchips. Furthermore, we did not observe any deleterious effects with low-conductivity buffers. We investigated the rapid and highly sensitive detection of mycoplasma contamination and the real samples of circulating cell-free DNA (cfDNA), which gave a limit of detection (LOD) as low as 2.3 ng mL−1. Owing to the significant reduction in cost, ease of operation, and fast separation capabilities demonstrated in this work, MCE can be a viable alternative to the usual slab gel electrophoresis running in most biological laboratories.

B-PO01-060 THE DELIVERY PATTERN OF RADIOFREQUENCY APPLICATIONS IN A LEFT ATRIAL-ESOPHAGEAL ABLATION MODEL DEMONSTRATES A HEAT STACK EFFECT: IMPLICATIONS FOR LEFT ATRIAL POSTERIOR WALL ABLATION

Effective radiofrequency (RF) applications along the posterior wall of the left atrium can result in esophageal (eso) thermal injury. Tissue heat capacitance may lead to accumulation of RF induced heating from contiguous lesions and cause additive heating, a phenomenon known as heat stacking. The delivery of interrupted compared to contiguous lesions may minimize heat stacking and reduce luminal eso heating. To evaluate the impact of RF application pattern and power on eso heat stacking in an in vitro model. A gel slab was created from Phytagel, deionized water and sodium chloride with a target impedance of 150 Ω. An eso temperature probe was placed under the gel in a saline bath. Temperature was recorded during delivery of 8 lesions in either a contiguous pattern or non-contiguous pattern. Lesions were delivered with an irrigated catheter with high power short duration (HPSD, 45 watts, 8 seconds) or low power longer duration (LPLD, 30 watts, 20 seconds) with a target force of 10 g. Tabled 1HPSDLPLDUninterrupted contiguousInterrupted non-contiguousUninterrupted contiguousInterrupted non-contiguousmean delta impedance (ohms)-36 +/- 4.3-11.1 +/- 1.2p < 0.01-31 +/-7.1-10.1 +/- 2.0p < 0.01mean force (grams)9.8 +/- 1.514 +/- 1.6p < 0.018.8 +/- 1.614 +/- 1.6p < 0.01mean time (seconds)64 +/- 086 +/- 7.2p < 0.01160 +/- 0188 +/- 6p < 0.01mean delta Temp (C)4.5 +/- 0.732.0 +/- 0.78p < 0.018.2 +/- 1.02.7 +/- 0.99p < 0.01 Open table in a new tab Absolute temperature change during uninterrupted contiguous were greater compared to interrupted non-contiguous. LPLD lesions had a greater temperature change than HPSD lesions (uninterrupted contiguous p < 0.01; interrupted non-contiguous P = 0.09). Lesion delivery pattern is an important cause for heat stacking and increased esophageal temperatures. Lesions delivered with LPLD had greater heat stacking effect compared to HPSD, which may reflect a greater amount of conductive heat transfer.

Aloe vera Gel Drying by Refractance Window®: Drying Kinetics and High-Quality Retention.

In most cases, conventional drying produces inferior quality products and requires higher drying times. A continuous pilot Refractance Window® equipment was used to dry Aloe vera gel slabs of 5 and 10 mm thick at 60, 70, 80, and 90 °C, seeking a dry product with high-quality retention. Based on five empirical models, drying kinetics, diffusion coefficient, and activation energy were analyzed. Midilli–Kuck was the best predicting model. Short drying times (55–270 min) were needed to reach 0.10 g water/g solid. In addition, the technique yielded samples with high rehydration capacity (24–29 g water/g solid); high retention of color (∆E, 3.74–4.39); relatively low losses of vitamin C (37–59%) and vitamin E (28–37%). Regardless of the condition of temperature and sample thickness, a high-quality dried Aloe vera gel could be obtained. Compared with other methods, Refractance Window® drying of Aloe vera achieved shorter drying times with higher quality retention in terms of color, vitamins C and E, and rehydration. Finally, the dried Aloe vera gel could be reconstituted to a gel close to its fresh state by rehydration.

A comparative study of CE-SDS, SDS-PAGE, and Simple Western-Precision, repeatability, and apparent molecular mass shifts by glycosylation.

SDS gel electrophoresis is a commonly used approach for monitoring purity and apparent molecular mass (Mr) of proteins, especially in the field of quality control of biopharmaceutical proteins. The technological installation of CE-SDS as the replacement of the slab gel technique (SDS-PAGE) is still in progress, leading to a continuous improvement of CE-SDS instruments. Various CE-SDS instruments, namely Maurice (CE-SDS/CE-SDS PLUS) and Wes by ProteinSimple as well as the microchip gel electrophoresis system LabChip® GXII Touch™ HT by PerkinElmer were tested for precision and repeatability compared to SDS-PAGE (Bio-Rad). For assessing these quality control parameters, standard model proteins with minor post-translational modifications were used. Overall, it can be concluded that the CE-SDS-based methods are similar to SDS-PAGE with respect to these parameters. Quality characteristics of test systems gain more significance by testing proteins that do not behave like model proteins. Therefore, glycosylated proteins were analyzed to comparatively investigate the influence of glycosylation on Mr determination in the different instruments. In some cases, high deviations were found both among the methods and with regard to reference values. This article provides possible explanations for these findings.

Sensitivity and applications of the PCR Single-Strand Conformation Polymorphism method.

PCR Single-Strand Conformation Polymorphism is a method used to identify and detect mutations and is now well known for its many applications on living beings. This paper will discuss the experimental details, limitations and sensitivity of the PCR Single-Strand Conformation Polymorphism method in relation to all existing literature available to us until today. Genomic DNA extraction, PCR amplification and Single-Strand Conformation Polymorphism conditions (concentration of polyacrylamide slab gel electrophoresis, dissociation treatment of double- stranded DNA) and comparison with PCR Restriction Fragment Length Polymorphism are presented. Since its discovery in 1989, there have been many variations, innovations, and modifications of the method, which makes it very easy, safe, fast and for this reason widely applied in clinical diagnostic, forensic medicine, biochemical, veterinary, microbiological, food and environmental laboratories. One of the possible applications of the method is the diagnosis and identification of mutations in new strains of coronaviruses, because science needs more tools to tackle the problem of this pandemic. The PCR Single-Strand Conformation Polymorphism method can be applied in many cases provided that control samples are available and the required conditions of the method are achieved.