Application of Liquid-Phase Preparative Isoelectric Focusing for Comparative Analysis of Abundancy in Differentially Extracted Bacterial Protein of S.Typhi and Invasive Non-Typhoidal Salmonella

Abstract

Isolation, purification, and separation of complex mixtures is crucial in proteomic research. The conventional electrophoresis method for antigen characterization has limitations in separating low abundance components and selectively enriching important proteins with high degree of purity. Two-dimensional gel electrophoresis was introduced to overcome these limitations, but it also has inherent shortcomings in detecting hydrophobic proteins, low abundance proteins, or samples with proteins of various concentrations. Therefore, this study aims to develop an innovative approach for the enrichment and characterization of immunoreactive components found in differentially extracted whole cell bacterial protein derived from S.Typhi and S.spp. A modified liquid phase preparative isoelectric focusing (IEF) and SDS-PAGE method was used to purify and characterize the proteins. The modified liquid phase IEF efficiently fractionated the proteins into 20 fractions based on the pI value, providing a high-resolution power for protein separation, high throughput, and ease of performance. The fractionated proteins were then analysed by SDS-PAGE for their molecular weight, providing a simple and cost-efficient method for protein analysis. This innovative approach for the enrichment and characterization of immunoreactive components in differentially extracted whole cell bacterial protein derived from S.Typhi and S.spp has the potential to revolutionize the diagnosis and treatment of typhoid fever and other related diseases. By improving the sensitivity and accuracy of protein analysis, this study may lead to identification of exclusive disease biomarkers for early, accurate diagnosis of diseases, improved prognosis and treatment outcomes.