Interrogation of an autofluorescence-based method for protein fingerprinting

Siddaramaiah, Manjunath and Rao, Satish BS and Joshi, Manjunath B and Datta, Anirbit and Chandra, Subhash and Nayak, Subramanya G and Satyamoorthy, K and Mahato, Krishna K (2018) Interrogation of an autofluorescence-based method for protein fingerprinting. Journal of Biophotonics, 11. pp. 1-11. ISSN 1864-063X

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In the present study, we have designed a laser-induced fluorescence (LIF) based instrumentation and developed a sensitive methodology for the effective separation,visualization, identification and analysis of proteins on a single platform. In this method, intrinsic fluorescence spectra of proteins were detected after separation on 1 or 2 dimensional Sodium Dodecyl Sulfate–Tris(2-carboxyethyl)phosphine (SDS-TCEP) polyacrylamide gel electrophoresis (PAGE) and the data were analyzed. The MATLAB assisted software was designed for the development of PAGE fingerprint for the visualization of protein after 1- and 2-dimensional protein separation. These provided objective parameters of intrinsic fluorescence intensity, emission peak, molecular weight and isoelectric point using a single platform. Further, the current architecture could differentiate the overlapping proteins in the PAGE gels which otherwise were not identifiable by conventional staining, imaging and tagging methods. Categorization of the proteins based on the presence or absence of tyrosine or tryptophan residues and assigning the corresponding emission peaks (309-356 nm) with pseudo colors allowed the detection of proportion of proteins within the given spectrum. The present methodology doesn't use stains or tags, hence amenable to couple with mass spectroscopic measurements. This technique may have relevance in the field of proteomics that is used for innumerable applications.

Item Type: Article
Uncontrolled Keywords: 1D/2D polyacrylamide gel electrophoresis (PAGE); Autofluorescence; Mass spectrometry; Polyacrylamide gel electrophoresis (PAGE); Protein fingerprint; Tris(2-corboxyethyl)phosphine (TCEP)
Subjects: Life Sciences > MLSC Manipal
Depositing User: KMC Library
Date Deposited: 03 Jan 2019 10:23
Last Modified: 03 Jan 2019 10:23

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