scholarly journals On the Applicability of Electrophoresis for Protein Quantification

Polymers ◽  
2021 ◽  
Vol 13 (22) ◽  
pp. 3971
Author(s):  
Karina Dome ◽  
Zoya Akimenko ◽  
Aleksey Bychkov ◽  
Yuri Kalambet ◽  
Oleg Lomovsky
Keyword(s):  
Gel Electrophoresis ◽  
Protein Denaturation ◽  
Methodological Approach ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein Quantification ◽  
Electrophoretic Data ◽  
Model Protein ◽  
World Protein ◽  
Mathematical Processing ◽  
Mathematical Data Processing

Polyacrylamide gel electrophoresis 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. This paper shows the feasibility of routine digital image acquisition and mathematical processing of electropherograms for protein quantification when using vertical gel electrophoresis and Chrom & Spec software. 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 the 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 as an affordable, convenient and quick tool.

scholarly journals On the Applicability of Electrophoresis for Protein Quantification

2021 ◽  
Author(s):  
Karina Dome ◽  
Zoya Akimenko ◽  
Aleksey Bychkov ◽  
Yuri Kalambet ◽  
Oleg Lomovsky
Keyword(s):  
Qualitative Method ◽  
Protein Denaturation ◽  
Methodological Approach ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein Quantification ◽  
Electrophoretic Data ◽  
Model Protein ◽  
World Protein ◽  
Mathematical Processing ◽  
Mathematical Data Processing

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.

scholarly journals Protein Transfer in Mainstream and Sidestream Cigarette Smoke

2004 ◽  
Vol 21 (1) ◽  
pp. 9-14
Author(s):  
R Voisine ◽  
F Côté ◽  
J Verreault ◽  
A Porter
Keyword(s):  
Cigarette Smoke ◽  
Tobacco Smoke ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Solvent System ◽  
Protein Transfer ◽  
Western Immunoblotting ◽  
Sidestream Smoke ◽  
Sds Page ◽  
Model Protein

AbstractProtein transfer in tobacco smoke has been studied using the protease, Savinase™, as a model protein. Mainstream and sidestream smoke were collected from cigarettes to which Savinase had been added at various concentrations. Savinase was extracted from the smoke condensate with an organic solvent system before being precipitated and further identified by denaturing polyacrylamide gel electrophoresis (SDS-PAGE) and Western immunoblotting. The detection limit of the method, based on addition of Savinase to the smoke condensate, was 25 µg in mainstream and 100 µg in sidestream smoke. At a Savinase concentration of 6000 µg per gram of tobacco, the methodology allows the detection of protein transfer as low as 0.009% and 0.054% in mainstream and sidestream smoke, respectively. Using this approach, it was shown that there is no detectable Savinase in the mainstream and sidestream smoke of filtered and unfiltered cigarettes containing up to 6000 µg of Savinase per gram of tobacco. These facts strongly suggest that there is no significant transfer of protein from tobacco into cigarette smoke.

Sodium dodecyl sulfate polyacrylamide gel electrophoresis of serum after protein denaturation in the presence or absence of 2-mercaptoethanol.

1984 ◽  
Vol 30 (3) ◽  
pp. 475-479 ◽  
Author(s):  
T Marshall
Keyword(s):  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Serum Proteins ◽  
Protein Denaturation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Major Protein ◽  
Dodecyl Sulfate ◽  
Electrophoretic Techniques

Abstract Human serum proteins were evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis after protein denaturation in the presence or absence of 2-mercaptoethanol. Both electrophoretic techniques give characteristic and reproducible banding patterns and achieve a high degree of resolution within the limits of a one-dimensional separation. The major protein bands have been identified, and the merits of the two techniques are compared. Protein denaturation in the absence of 2-mercaptoethanol gives more discrete bands for the purpose of protein identification by maintaining the disulfide-bridge-dependent polypeptide associations characteristic of many serum proteins. However, simultaneous use of both electrophoretic techniques enhances identification by exploiting the response of an individual protein to mercaptoethanol treatment. The clinical potential of this approach for detecting protein disorders is discussed.

The quantitative measurement of whey proteins using polyacrylamide-gel electrophoresis

1976 ◽  
Vol 43 (2) ◽  
pp. 259-265 ◽  
Author(s):  
Robyn M. Hillier
Keyword(s):  
Gel Electrophoresis ◽  
Protein Denaturation ◽  
Quantitative Estimation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Whey Proteins ◽  
Skim Milk ◽  
Internal Standard ◽  
Polyacrylamide Gel ◽  
Native Whey ◽  
Β Lactoglobulin

SummaryA method is described for the quantitative estimation of a mixture of whey proteins by spectrophotometric scanning of the stained protein bands following polyacrylamide-gel electrophoresis. The incorporation of β-lactoglobulin A as an internal standard improves the accuracy of the technique. The method has been used to measure residual native whey proteins in milk after heating. Results are presented for whey protein denaturation in skim-milk after heat treatment at 130 and 140 °C for various periods of time.

104 ESTROUS CYCLE-DEPENDENT CHANGES IN THE BOVINE ENDOMETRIUM PROTEOME

2009 ◽  
Vol 21 (1) ◽  
pp. 152
Author(s):  
T. Fröhlich ◽  
R. Kashirin ◽  
P. Bolbrinker ◽  
H.-D. Reichenbach ◽  
E. Wolf ◽  
...  
Keyword(s):  
Corpus Luteum ◽  
Estrous Cycle ◽  
Gel Electrophoresis ◽  
Hormonal Regulation ◽  
Molecular Mechanisms ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein Quantification ◽  
Progesterone Level ◽  
Uterine Receptivity ◽  
Ph Gradients

Among the reproductive tissues, the endometrium plays a central role in the context of embryo-maternal communication and pregnancy recognition. During the estrous cycle, characteristic morphological and functional changes occur in the bovine endometrium, being crucial for uterine receptivity. These changes are mainly regulated by the hormones progesterone, estradiol, and oxytocin. The bovine estrous cycle, with a length of 21 days, can be divided into 4 stages: i) estrus (considered as Day 0, low progesterone level and time of ovulation); ii) metestrus (Days 1 to 5, corpus luteum formation, rising progesterone level); iii) diestrus (Days 6 to 17, high progesterone); and iv) proestrus (Days 18 to 20, corpus luteum degeneration; declining progesterone). The principles of hormonal regulation during the estrous cycle are well understood; however, in-depth knowledge of the detailed molecular mechanisms is still incomplete. To elucidate the underlying biochemical processes, the proteomes of bovine endometrial samples of all 4 stages (cycle Day 0, Day 3.5, Day 12, and Day 18) were compared in a quantitative manner. To maximize the accuracy of protein quantification, sophisticated 2-dimensional (2D) fluorescence difference gel electrophoresis (2D-DIGE) experiments were performed, using internal pooled standards for inter-gel normalization. To enhance the resolution of 2D-polyacrylamide gel electrophoresis separation, the proteins were analyzed by 2 overlapping pH gradients. In total, 28 individual DIGE experiments (14 2D gels × 2 pH gradients) were performed, corresponding to 84 gel images. With a refined statistical analysis of spot intensities, we were able to identify a total of 91 spots altered by at least a factor of ±2 (P < 0.05) in intensity between at least 2 of the 4 stages. Matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) identification of these spots showed that they originated from 66 different proteins. Moreover, for 14 of these proteins, several polymorphic variants could be identified. Gene ontology analysis of the protein IDs revealed a broad diversity of biological and biochemical functions as well as cellular localizations of these proteins. Several proteins detected (e.g. FK506 and 20 alpha-HSD) are crucial components for uterine receptivity and represent interesting targets for further functional studies. This study was supported by the Deutsche Forschungsgemeinschaft (FOR 478).

Ultrastructural localization of specific nucleoprotein fractions

1978 ◽  
Vol 36 (2) ◽  
pp. 204-205
Author(s):  
G. L. Brown
Keyword(s):  
Gel Electrophoresis ◽  
Mouse Liver ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ultrastructural Localization ◽  
Polyacrylamide Gel ◽  
Acid Extraction ◽  
Acid Solutions ◽  
Low Salt ◽  
Liver Nuclei ◽  
Phosphate Groups

Bismuth (Bi) stains nucleoproteins (NPs) by interacting with available amino and primary phosphate groups. These two staining mechanisms are distinguishable by glutaraldehyde crosslinking (Fig. 1,2).Isolated mouse liver nuclei, extracted with salt and acid solutions, fixed in either formaldehyde (form.) or gl utaraldehyde (glut.) and stained with Bi, were viewed to determine the effect of the extractions on Bi stainina. Solubilized NPs were analyzed by SDS-polyacrylamide gel electrophoresis.Extraction with 0.14 M salt does not change the Bi staining characteristics (Fig. 3). 0.34 M salt reduces nucleolar (Nu) staining but has no effect on interchromatinic (IC) staining (Fig. 4). Proteins responsible for Nu and glut.- insensitive IC staining are removed when nuclei are extracted with 0.6 M salt (Fig. 5, 6). Low salt and acid extraction prevents Bi-Nu staining but has no effect on IC staining (Fig. 7). When nuclei are extracted with 0.6 M salt followed by low salt and acid, all Bi-staining components are removed (Fig. 8).

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