Bradford protein assay

is also catalysed in the other direction by LDH during the Cori cycle. This reaction can be assayed using spectrophotometric techniques; peak absorbance of NADH is at 340nm, but the peak absorbance of NAD+ is 259nm (Powers, et al., 2007). Enzyme assays are performed to measure the rate of an enzyme-catalysed reaction, known as the enzyme activity. Many factors can affect the enzyme activity (including temperature, pH, substrate concentration), so all conditions apart from the one being…

natively folded proteins are not molecular weight dependent, the proteins would be traveling at different speeds in an electric field. In order to separate the proteins based on their molecular weight, the tertiary structure of the protein needs to be destroyed making the protein a linear molecule. Sodium dodecyl sulfate (SDS) plays an important role in this mechanism. Sodium dodecyl sulfate is an amphipathic detergent whose role is to break down the tertiary structure of a protein making it a…

Rabia Iqbal Biol 395-1005 lab 01/31/2015 Broadford assay Absorbance Figure 1. The concentration and absorbance of Bradford assay. Four samples containing Bovine serum albumin, diluent, and Bradford. At 0.25 mg/ml concentration the absorbance was 0.109 nm and the relation between absorbance and the concentration held constant throughout except at the 0.50 mg/ml when the concentration absorbance was 0.186 nm. Figure showed that as the concentration is increased the absorbance elevates also. All…

make sure it stays below 20 C. This was done until the solution was homogeneous. The solution was then transferred to a centrifuge tube and centrifuged. The supernatant was aliquoted and stored in a -20 C for use in future experiments. Enzymatic Assay of LDH by Spectrophotometry. One tube of tissue extract was thawed out on ice and a…

enzyme will be eluted at high pH. The Bradford assay will be used to determine the protein concentration and the specific activity of the solutions containing the enzyme. Size-exclusion chromatography separates molecules…

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For each group cDNA was synthesized from 500 ng of RNA using Maxima Universal First Strand cDNA Synthesis kit (Thermo Scientific, catalogue no-K1661) and consecutively 50 ng of cDNA was used for quantitative real time PCR assay. cDNA was amplified in a thermal cycle (CFX96 PCR system, BioRadLaboretories) in 20 µl reaction mixture containing 0.5 µl cDNA template (50 ng), 12.5 µl Maxima SYBR green/fluorescent qPCR master mix (2X, Thermo Scientific), 0.3 µl forward and reverse…

collected. Protein Extraction and Quantification The absorbance vs concentration curve prepared from the Bradford Protein Assay standards was found to have a linear regression of y = 2.2184x + 0.0626 as shown in Figure 1. The absorbances of the liver, muscle, and adipose tissue were used to determine the protein concentrations of the samples. The percent protein was found to be 15.64% in liver tissue, 4.13% in muscle…

was used as crude enzyme in subsequent studies. The protein concentration in the enzyme preparation was determined following the method of Bradford…

Analysis of Consumer Products with Unknown Protein Concentrations Using Spectrophotometry Abstract Consumer products typically give detailed information about their ingredients and nutrients. However, the concentration of a specific nutrient may not always be clear from just the label information alone, and in some cases the product labels may not be accurate. Thus, it may be hard to compare the concentration level of a certain nutrient, such as protein, amongst different products. In this…