Comparative Analysis of Electrophoresis and Interferometric Optical Detection Method for Molecular Weight Determination of Proteins

Overview

Journal Heliyon

Specialty Social Sciences

Date 2024 Sep 4

PMID 39229532

Authors

B Santamaria

A L Hernandez

M F Laguna

M Holgado

Affiliations

Soon will be listed here.

Abstract

Analytical detection methods play a pivotal role in scientific research, enabling the identification and quantification of specific analytes in various disciplines. This scientific report aims to compare two very different methodologies for determining the Molecular Mass (MM, also known as Molecular Weight, MW) of proteins: electrophoresis gel and the Interferometric Optical Detection Method (IODM). For this purpose, several proteins with different MM were selected. The electrophoresis technique was employed to validate the structure and MM of different parts or fragments of the Matrix Metallopeptidase 9 antibody (anti-MMP9), antibody against S100 calcium binding protein A6 (anti-S100A6) and Cystatin S4 antibody (anti-CST4) by examining the presence of bands with expected sizes. The IODM was applied to study the above-mentioned proteins (part of the antibodies) together with the protein G, as a reference to correlate the MM and protein sizes with the measured signal. We report the evidence of IODM as a competitive analytical approach for the determination of the MM of proteins for the first time. This innovative method allows for accurate MM determination using minimal sample volumes and concentrations, employing a simple experimental procedure that eliminates the requirement for protein denaturation.

References

Erickson H . Size and shape of protein molecules at the nanometer level determined by sedimentation, gel filtration, and electron microscopy. Biol Proced Online. 2009; 11:32-51. PMC: 3055910. DOI: 10.1007/s12575-009-9008-x. View

Casquel R, Holgado M, Laguna M, Hernandez A, Santamaria B, Lavin A . Engineering vertically interrogated interferometric sensors for optical label-free biosensing. Anal Bioanal Chem. 2020; 412(14):3285-3297. PMC: 7214506. DOI: 10.1007/s00216-020-02411-3. View

Wiesner R, Scheller C, Krebs F, Watzig H, Oltmann-Norden I . A comparative study of CE-SDS, SDS-PAGE, and Simple Western: Influences of sample preparation on molecular weight determination of proteins. Electrophoresis. 2020; 42(3):206-218. DOI: 10.1002/elps.202000199. View

Kirley T, Norman A . Unfolding of IgG domains detected by non-reducing SDS-PAGE. Biochem Biophys Res Commun. 2018; 503(2):944-949. PMC: 6095737. DOI: 10.1016/j.bbrc.2018.06.100. View

Laguna M, Holgado M, Sanza F, Lavin A, Lopez A, Casquel R . Optimization of dengue immunoassay by label-free interferometric optical detection method. Sensors (Basel). 2014; 14(4):6695-700. PMC: 4029634. DOI: 10.3390/s140406695. View

Vavricka S, Burri E, Beglinger C, Degen L, Manz M . Serum protein electrophoresis: an underused but very useful test. Digestion. 2009; 79(4):203-10. DOI: 10.1159/000212077. View

Goetz H, Kuschel M, Wulff T, Sauber C, Miller C, Fisher S . Comparison of selected analytical techniques for protein sizing, quantitation and molecular weight determination. J Biochem Biophys Methods. 2004; 60(3):281-93. DOI: 10.1016/j.jbbm.2004.01.007. View

Ofarrell P . High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975; 250(10):4007-21. PMC: 2874754. View

Chrambach A, Rodbard D . Polyacrylamide gel electrophoresis. Science. 1971; 172(3982):440-51. DOI: 10.1126/science.172.3982.440. View

10.

Escuder-Gilabert L, Bermudez-Saldana J, Villanueva-Camanas R, Medina-Hernandez M, Sagrado S . Reliability of the retention factor estimations in liquid chromatography. J Chromatogr A. 2004; 1033(2):247-55. DOI: 10.1016/j.chroma.2004.01.038. View

11.

Holgado M, Sanza F, Lopez A, Lavin A, Casquel R, Laguna M . Description of an advantageous optical label-free biosensing interferometric read-out method to measure biological species. Sensors (Basel). 2014; 14(2):3675-89. PMC: 3958242. DOI: 10.3390/s140203675. View