![]() ![]() Two-dimensional gel electrophoresis (2-DE) represents one of the main technologies for quantitative proteomics with advantages and disadvantages. Quantification using two dimensional electrophoresis This system consists of the calibrated optical filter with very high angular resolution and the interaction of light with this crystal forms a resonance at a wavelength which correlates to concentration and refractive index near the crystal. An alternative method for label free protein quantification in clear liquid is cuvette-based SPR technique, that simultaneously measures the refractive index ranging 1.0 to 1.6 nD and concentration of the protein ranging from 0.5 µL to 2 mL in volume. Other more accurate spectrophotometric procedures for protein quantification include the Biuret, Lowry, BCA, and Bradford methods. This method is also inaccurate due to the possibility of nucleic acid contamination. However, this method is not the most accurate because the composition of proteins can vary greatly and this method would not be able to quantify proteins that do not contain the aforementioned amino acids. The concentration of a protein can be determined by measuring the OD at 280 nm on a spectrophotometer, which can be used with a standard curve assay to quantify the presence of tryptophan, tyrosine, and phenylalanine. The concentration of a certain protein in a sample may be determined using spectrophotometric procedures. In contrast to 2-DE, which requires MS for the downstream protein identification, MS technology can identify and quantify the changes. However, a recent developed method of quantitative dot blot (QDB) analysis is able to measure both the absolute and relative quantity of an individual proteins in the sample in high throughput format, thus open a new direction for proteomic research. Quantitative proteomics is mainly performed by two-dimensional gel electrophoresis (2-DE), preparative one-dimensional gel electrophoresis, or mass spectrometry (MS). For example, this approach can be used to compare samples from healthy and diseased patients. Rather than just providing lists of proteins identified in a certain sample, quantitative proteomics yields information about the physiological differences between two biological samples. qualitative) proteomics, but include quantification as an additional dimension. The methods for protein identification are identical to those used in general (i.e. Finally, these results allowed to reinforce the hypothesis of oxidative damage in erythrocyte membrane proteins as molecular mechanism of human adaptation to malaria infection.Quantitative proteomics is an analytical chemistry technique for determining the amount of proteins in a sample. Therefore, both polymorphisms promote carbonylation on the same membrane proteins. Erythrocytes with G6PD deficient and SCT showed higher carbonyl index values than control and similar profiles of carbonylated proteins moreover, cytoskeletal and stress response proteins were identified as the main targets of oxidative damage. Besides, protein carbonylation profiles were obtained by Western blot and complemented with mass spectrometry using MALDI-TOF-TOF analysis. Carbonyl index by dot blot assay was used to calculate the variation of oxidative damage during the asexual phases. falciparum 3D7 were used to quantify oxidative damage in membrane proteins of erythrocytes with G6PD deficient and SCT. Here, synchronous cultures at high parasitemia levels of P. Nevertheless, mechanisms are not completely understood at molecular level for each polymorphism yet, and even less if are commons for several of them. There exist in both a prooxidant environment that favors the oxidative damage on membrane proteins, which probably is part of molecular protector mechanisms. Both are considered the result of the selective pressure exerted by Plasmodium parasites over human genome, due to a certain degree of resistance to the clinical symptoms of severe malaria. Deficiency of glucose-6-phosphate dehydrogenase (G6PD) and sickle cell trait (SCT) are described as the polymorphic disorders prevalent in erythrocytes. ![]()
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