Quantification of Soluble or Insoluble Fractions of Leishmania Parasite Proteins in Microvolume Applications: A Simplification to Standard Lowry Assay. (13th March 2020)

• Record Type: Journal Article
• Authors: Deepachandi, Bhagya; Weerasinghe, Sudath; Andrahennadi, Thisira Priyantha; Karunaweera, Nadira D.; Wickramarachchi, Nadeeja; Soysa, Preethi; Siriwardana, Yamuna
• Other Names: Abdel-Rehim Mohamed Academic Editor.
• Is Part Of: International journal of analytical chemistry. Volume 2020(2020)
• Publication Date: 2020-03-13
• Subjects: Chemistry, Analytic -- Periodicals; 543.05
• DOI: 10.1155/2020/6129132
• Languages: English
• ISSNs: 1687-8760

Abstract

Abstract : Protein quantification is often an essential step in any research field that involves proteins. Although the standard Lowry assay and its modifications are most abundantly used in protein quantification, the existing methods are rigid or often demonstrate nonlinearity between protein concentration and color intensity. A method for fast and accurate qualitative and/or quantitative determination of total soluble/insoluble proteins or micro-well plate immobilized proteins isolated from Leishmania parasites in microvolumes was described in the current study. Improvements in cost-effective techniques are necessary to increase the research outputs in resource-limited settings. This method is a modification to the established Lowry assay for protein quantification. Concentrations of unknown samples were calculated using a standard curve prepared using a standard series of bovine serum albumin (BSA). The optimized reagents were 2 N NaOH (sodium hydroxide), 2% Na2 CO3 (sodium carbonate), 1% CuSO4 (copper sulfate), 2% KNaC4 H4 O6 (potassium sodium tartrate), and 2 N Folin and Ciocalteu's phenol. This modified protein assay was sensitive for quantifying Leishmania proteins in a total crude extract or in a soluble fraction within the approximate range of 10–500 μ g/ml (1–50 μ g/assay) and showed a linearity between color intensity and concentration of the protein. This is an easier, fast, and accurate method for quantifying proteins with microvolumes in a cost-effective manner for routine use in research laboratories in resource-limited settings.