Time for Multiple Extraction Methods in Proteomics? A Comparison of Three Protein Extraction Methods in the Eustigmatophyte Alga Microchloropsis gaditana CCMP526

2017 ◽  
Vol 21 (11) ◽  
pp. 678-683 ◽  
Author(s):  
Anbarasu Karthikaichamy ◽  
Pranali Deore ◽  
Vineeta Rai ◽  
Dieter Bulach ◽  
John Beardall ◽  
...  
Keyword(s):  
Protein Extraction ◽  
Extraction Methods

scholarly journals Optimization of Protein Extraction Method for 2DE Proteomics of Goat’s Milk

Molecules ◽  
2020 ◽  
Vol 25 (11) ◽  
pp. 2625
Author(s):  
Muzammeer Mansor ◽  
Jameel R. Al-Obaidi ◽  
Nurain Nadiah Jaafar ◽  
Intan Hakimah Ismail ◽  
Atiqah Farah Zakaria ◽  
...  
Keyword(s):  
Extraction Method ◽  
Protein Extraction ◽  
Chloroform Solution ◽  
Milk Proteins ◽  
Peptide Mass Fingerprinting ◽  
Extraction Methods ◽  
Dairy Goats ◽  
Goat’S Milk ◽  
Peptide Mass ◽  
Goat's Milk

Two-dimensional electrophoretic (2DE)-based proteomics remains a powerful tool for allergenomic analysis of goat’s milk but requires effective extraction of proteins to accurately profile the overall causative allergens. However, there are several current issues with goat’s milk allergenomic analysis, and among these are the absence of established standardized extraction method for goat’s milk proteomes and the complexity of goat’s milk matrix that may hamper the efficacy of protein extraction. This study aimed to evaluate the efficacies of three different protein extraction methods, qualitatively and quantitatively, for the 2DE-proteomics, using milk from two commercial dairy goats in Malaysia, Saanen, and Jamnapari. Goat’s milk samples from both breeds were extracted by using three different methods: a milk dilution in urea/thiourea based buffer (Method A), a triphasic separation protocol in methanol/chloroform solution (Method B), and a dilution in sulfite-based buffer (Method C). The efficacies of the extraction methods were assessed further by performing the protein concentration assay and 1D and 2D SDS-PAGE profiling, as well as identifying proteins by MALDI-TOF/TOF MS/MS. The results showed that method A recovered the highest amount of proteins (72.68% for Saanen and 71.25% for Jamnapari) and produced the highest number of protein spots (199 ± 16.1 and 267 ± 10.6 total spots for Saanen and Jamnapari, respectively) with superior gel resolution and minimal streaking. Six milk protein spots from both breeds were identified based on the positive peptide mass fingerprinting matches with ruminant milk proteins from public databases, using the Mascot software. These results attest to the fitness of the optimized protein extraction protocol, method A, for 2DE proteomic and future allergenomic analysis of the goat’s milk.

Comparison of Two Protein Extraction Methods for Tea Residues

2011 ◽  
Vol 138-139 ◽  
pp. 933-936 ◽  
Author(s):  
Xuan Chen ◽  
Hong Yu Luo ◽  
Jun Yu ◽  
Peng Xiang Yue ◽  
Lin Zhou ◽  
...  
Keyword(s):  
High Temperature ◽  
Protein Extraction ◽  
Ethanol Concentration ◽  
Extraction Methods ◽  
Extraction Yield ◽  
Factorial Experiments ◽  
Extraction Technology ◽  
Digestion Method ◽  
Solid Ratio ◽  
Lower Temperature

Alcohol-alkali method and base digestion method were investigated to extract proteins from tea residues, respectively. According to single factorial experiments, results showed that the optimal extraction technology of alcohol-alkali method were pH 12, temperature of 80 °C, ethanol concentration of 60%, liquid-solid ratio of 40, 60 min, and the protein extraction rate reached 15.0%. And the optimal extract conditions of base digestion were pH 12, temperature of 80 °C, liquid-solid ratio of 50, 80 min, which made the protein yield reached 31.5%. Furthermore, alcohol-alkali method was more beneficial to protein extraction from tea residues under lower temperature and weak alkali condition (40-60 °C, pH 8-10). While base digestion had higher extraction yield under high temperature and strong alkali condition (60-80 °C, pH 11-12).

scholarly journals Evaluation of effective protein extraction procedure to profile petiole of Moringa oleifera

2020 ◽  
Vol 7 (2) ◽  
pp. 214
Author(s):  
Zetty Amirah Zulkifli ◽  
Zaidah Rahmat
Keyword(s):  
Moringa Oleifera ◽  
Protein Extraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Extraction Procedure ◽  
Electrophoretic Pattern ◽  
Antioxidant Property ◽  
Extraction Methods ◽  
Protein Profiling ◽  
Sds Page

Moringa oleifera is widely known as multipurpose tree since all of its parts confer multiple functions. The leaf is highly favourable among consumers while the petiole is mostly wasted. There are numerous studies on the flavonoid and antioxidant property of the stem and twig. However, study on the petiole has never been done. There-upon, this study was conducted to develop protein profiling of the petiole. In this study, 6 different protein extraction methods were tested on the fresh petiole before its protein quantity and quality were checked via Bradford assay and Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE) respectively. The in-solution digestion was then done prior to LC-MS/MS analysis. The protein electrophoretic pattern from the SDS-PAGE proves that method 6 using Tris HCl buffer with incorporation of dithiothreitol (DTT) and phenylmethylsulfonyl fluoride (PMSF) confers the best quality of protein. It produced the highest number of visible individual bands compared to other methods. Meanwhile, 93 proteins were successfully identified via LCMS analysis where the protein, signal response and carbohydrate metabolism categories confer the highest percentage. High quality and content of the protein extracted from the petiole including the antioxidant, anticancer and antidiabetic protein identified suggested that consuming this part of the plant could enhance nutrients of human body.

Development of Efficient Protein Extraction Methods for Shotgun Proteome Analysis of Formalin-Fixed Tissues

2007 ◽  
Vol 6 (3) ◽  
pp. 1038-1047 ◽  
Author(s):  
Xiaogang Jiang ◽  
Xinning Jiang ◽  
Shun Feng ◽  
Ruijun Tian ◽  
Mingliang Ye ◽  
...  
Keyword(s):  
Protein Extraction ◽  
Proteome Analysis ◽  
Extraction Methods ◽  
Formalin Fixed

scholarly journals Comparison of Conventional and Sustainable Lipid Extraction Methods for the Production of Oil and Protein Isolate from Edible Insect Meal

Foods ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 572 ◽  
Author(s):  
Laroche ◽  
Perreault ◽  
Marciniak ◽  
Gravel ◽  
Chamberland ◽  
...  
Keyword(s):  
Supercritical Co2 ◽  
Protein Extraction ◽  
Lipid Extraction ◽  
Tenebrio Molitor ◽  
Extraction Methods ◽  
Extraction Yield ◽  
Acheta Domesticus ◽  
Human Consumption ◽  
Alternative Source ◽  
Edible Insect

Edible insects represent an interesting alternative source of protein for human consumption but the main hurdle facing the edible insect sector is low consumer acceptance. However, increased acceptance is anticipated when insects are incorporated as a processed ingredient, such as protein-rich powder, rather than presented whole. To produce edible insect fractions with high protein content, a defatting step is necessary. This study investigated the effects of six defatting methods (conventional solvents, three-phase partitioning, and supercritical CO2) on lipid extraction yield, fatty profiles, and protein extraction and purification of house cricket (Acheta domesticus) and mealworm (Tenebrio molitor) meals. Ethanol increased the lipid extraction yield (22.7%–28.8%), irrespective of the insect meal used or the extraction method applied. Supercritical CO2 gave similar lipid extraction yields as conventional methods for Tenebrio molitor (T. molitor) (22.1%) but was less efficient for Acheta domesticus (A. domesticus) (11.9%). The protein extraction yield ranged from 12.4% to 38.9% for A. domesticus, and from 11.9% to 39.3% for T. molitor, whereas purification rates ranged from 58.3% to 78.5% for A. domesticus and from 48.7% to 75.4% for T. molitor.

scholarly journals Comparison of Different Protein Extraction Methods for Gel-Based Proteomic Analysis of Ganoderma spp.

2016 ◽  
Vol 35 (2) ◽  
pp. 100-106 ◽  
Author(s):  
Jameel R. Al-Obaidi ◽  
Noor Baity Saidi ◽  
Siti Rokhiyah Ahmad Usuldin ◽  
Siti Nahdatul Isnaini Said Hussin ◽  
Noornabeela Md Yusoff ◽  
...  
Keyword(s):  
Proteomic Analysis ◽  
Protein Extraction ◽  
Extraction Methods

scholarly journals Modified TCA/acetone precipitation of plant proteins for proteomic analysis

2018 ◽  
Author(s):  
Liangjie Niu ◽  
Hang Zhang ◽  
Zhaokun Wu ◽  
Yibo Wang ◽  
Hui Liu ◽  
...  
Keyword(s):  
Proteomic Analysis ◽  
Protein Modification ◽  
Protein Extraction ◽  
Extended Period ◽  
Extraction Methods ◽  
Precipitation Method ◽  
Plant Proteins ◽  
Prolonged Incubation ◽  
Acetone Precipitation ◽  
Modified Method

AbstractProtein extracts obtained from cells or tissues often require removal of interfering substances for the preparation of high-quality protein samples in proteomic analysis. A number of protein extraction methods have been applied to various biological samples. TCA/acetone precipitation and phenol extraction, a common method of protein extraction, is thought to minimize protein degradation and activity of proteases as well as reduce contaminants like salts and polyphenols. However, the TCA/acetone precipitation method relies on the complete pulverization and repeated rinsing of tissue powder to remove the interfering substances, which is laborious and time-consuming. In addition, by prolonged incubation in TCA/acetone, the precipitated proteins are more difficult to re-dissolve. We have described a modified method of TCA/acetone precipitation of plant proteins for proteomic analysis. Proteins of cells or tissues were extracted using SDS-containing buffer, precipitated with equal volume of 20% TCA/acetone, and washed with acetone. Compared to classical TCA/acetone precipitation and simple acetone precipitation, this protocol generates comparable yields, spot numbers, and proteome profiling, but takes less time (ca. 45 min), thus avoiding excess protein modification and degradation after extended-period incubation in TCA/acetone or acetone. The modified TCA/acetone precipitation method is simple, fast, and suitable for proteomic analysis of various plant tissues in proteomic analysis.

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