A Ponceau S Staining-Based Dot Blot Assay for Rapid Protein Quantification of Biological Samples

Despite the availability of a wide range of commercial kits, protein quantification is often unreliable, especially for tissue-derived samples, leading to uneven loading in subsequent experiments. Here we show that the widely used Bicinchoninic Acid (BCA) assay tends to underestimate protein concentrations of tissue samples. We present a Ponceau S staining-based dot-blot assay as an alternative for protein quantification. This method is simple, rapid, more reliable than the BCA assay, compatible with biological samples lysed in RIPA or 2x SDS gel-loading buffer, and also inexpensive.

Zymography-types, advantages, uses and troubleshooting - A detailed review

There are many methods for the study and quantification of enzymes in the body systems. Off-late, the method of zymography has assumed immense importance in diagnostic medicine. It is a method where enzymes are visualized using the substrate conversion technique. The product of reaction appears, or the substrate disappears and the mechanisms specific for the recognition of this measure the biochemical reaction. This procedure has many advantages, the most important ones being providing both qualitative and measurable data, such as molecular zymography methods for visualising hydrolytic enzymes and differentiating among whole molecules, degradation and complexes. There are three main types of zymography, namely- In-Gel zymography (IGZ), In-Situ zymography (ISZ) and In-vivo zymography (IVZ). There are many variations of this technique like transfer 2D and reverse zymography. This method is mainly used to study the expression of matrix metalloproteinases. Alternate to the conventional zymography techniques have been suggested, like usage of the new Ponceau S staining protocol that provides significant benefits in terms of assay usability and cost reduction and is comparatively faster and easier. Mild alterations in the actual procedure of zymography can take care of minor issues arising during the procedure.

Comparison of advanced oxidation processes for degrading Ponceau S dye. Application of photo-Fenton process.

The major part of this work is devoted to the examination of the degradation of Ponceau S (PS) in aqueous medium by using the photo-Fenton process. The influence of pH of the medium, oxidant dose (H2O2), ferrous ion dose and the presence of inorganic ions such as bicarbonate, sulphate and nitrate on degradation kinetics was also analyzed. Before that, the efficiency of this process was compared to three other advanced oxidation processes (AOPs) such as H2O2/UV, Fenton (Fe2+/H2O2), and solar photo-Fenton. It was found that all dye degradation kinetics obeyed a pseudo-first order, and their apparent rate constants were represented by the ratios: kapp (H2O2/UV/Fe2+) =0.295 kapp (sunlight/Fe2+/H2O2) =0.141 kapp (Fe2+/H2O2) = 0.111 kapp (H2O2/UV) =0.031.Under reference conditions (0.06 mM of PS in ultrapure water), photo-Fenton oxidation using 0.06 mM of Fe2+, 1mM of H2O2 and the pH optimal value of 3 yielded more than 94.3% in only10 min of dye degradation with an apparent rate constant of 0.2951 min-1. Elsewhere, the addition of the salts in aqueous medium has, in general, an inhibition effect on the decolorization kinetics due to controlled ionic interactions of metals with inorganic ions. The most adverse effect on PS disappearance rate was observed when bicarbonate ions were present in the treated medium.