High-sensitivity staining of proteins for one- and two-dimensional gel electrophoresis using post migration covalent staining with a ruthenium fluorophore

Electrophoresis
Caroline TokarskiChristian Rolando

Abstract

This paper describes the use of a ruthenium complex ((bis(2,2'-bipyridine)-4'-methyl-4-carboxybipyridine-ruthenium-N-succidimyl ester-bis(hexafluorophosphate), abbreviated below as ASCQ_Ru) commercially available and chemically pure. This new ruthenium complex ASCQ_Ru brings an activated ester, allowing the selective acylation of amino acid side chain amines for the post migration staining of proteins separated in 1-DE and 2-DE. The protocol used is a simple three-step protocol fixing the proteins in the gel, staining and then washing, as no lengthy destaining step is required. First the critical staining step was optimized. Although in solution the best described pH for acylating proteins with this reagent is phosphate buffer at pH 7.0, we found that best medium for in-gel staining is unbuffered ACN/water solution (20/80 v/v). The two other steps are less critical and classical conditions are satisfactory: fixing with 7% acetic acid/10% ethanol solution and washing four times for 10 min with water. Sensitivity tests were performed using 1-DE on protein molecular weight markers. We obtained a higher sensitivity than SYPRO Ruby with a detection limit of 80 pg of protein per well. However, contrary to SYPRO Ruby, ASCQ_Ru exhibits...Continue Reading

References

Jul 1, 1980·Analytical Biochemistry·B R OakleyN R Morris
Mar 1, 1996·Analytical Chemistry·A ShevchenkoM Mann
May 3, 2001·Electrophoresis·J P MaloneG S Gerstenecker
May 14, 1963·Biochimica Et Biophysica Acta·S Fazekas de St GrothA Datyner

❮ Previous
Next ❯

Citations

Aug 24, 2007·Applied Microbiology and Biotechnology·Matthias BerthJörg Bernhardt
Jun 21, 2011·Journal of Chemical Biology·Victoria J GauciJens R Coorssen
Nov 17, 2010·Analytical Biochemistry·Caroline TokarskiChristian Rolando
Jun 1, 2012·Electrophoresis·Junli JiaMing Zhou
Feb 13, 2018·Nature Communications·Maria Anna RapsomanikiMaría Rodríguez Martínez
Jan 23, 2020·Dalton Transactions : an International Journal of Inorganic Chemistry·Ankita SarkarParna Gupta
May 23, 2007·BioDrugs : Clinical Immunotherapeutics, Biopharmaceuticals and Gene Therapy·Thomas ThieleAndreas Greinacher
Oct 11, 2013·Proteomes·Barbara DeracinoisYannis Karamanos
Mar 21, 2007·Journal of Proteome Research·Lauren R HarrisJens R Coorssen

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.