Ruthenium (II) tris‐bathophenanthroline disulfonate is well suitable for Tris‐Glycine PAGE but not for Bis‐Tris gels

Moebius, Jan; Denker, Katrin; Sickmann, Albert

doi:10.1002/pmic.200600642

Cited by 9 publications

(13 citation statements)

References 12 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In comparison to silver staining RuBPS fluorescent staining shows a higher linearity in signal intensity and is more reproducible. Furthermore, RuPBS is more sensitive than Coomassie and offers compatibility with MS [23,24]. …”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Malignant transformation in a defined genetic background: proteome changes displayed by 2D-PAGE

et al. 2010

Self Cite

View full text Add to dashboard Cite

BackgroundCancer arises from normal cells through the stepwise accumulation of genetic alterations. Cancer development can be studied by direct genetic manipulation within experimental models of tumorigenesis. Thereby, confusion by the genetic heterogeneity of patients can be circumvented. Moreover, identification of the critical changes that convert a pre-malignant cell into a metastatic, therapy resistant tumor cell, however, is one necessary step to develop effective and selective anti-cancer drugs. Thus, for the current study a cell culture model for malignant transformation was used: Primary human fibroblasts of the BJ strain were sequentially transduced with retroviral vectors encoding the genes for hTERT (cell line BJ-T), simian virus 40 early region (SV40 ER, cell line BJ-TE) and H-Ras V12 (cell line BJ-TER).ResultsThe stepwise malignant transformation of human fibroblasts was analyzed on the protein level by differential proteome analysis. We observed 39 regulated protein spots and therein identified 67 different proteins. The strongest change of spot patterns was detected due to integration of SV40 ER. Among the proteins being significantly regulated during the malignant transformation process well known proliferating cell nuclear antigen (PCNA) as well as the chaperones mitochondrial heat shock protein 75 kDa (TRAP-1) and heat shock protein HSP90 were identified. Moreover, we find out, that TRAP-1 is already up-regulated by means of SV40 ER expression instead of H-Ras V12. Furthermore Peroxiredoxin-6 (PRDX6), Annexin A2 (p36), Plasminogen activator inhibitor 2 (PAI-2) and Keratin type II cytoskeletal 7 (CK-7) were identified to be regulated. For some protein candidates we confirmed our 2D-PAGE results by Western Blot.ConclusionThese findings give further hints for intriguing interactions between the p16-RB pathway, the mitochondrial chaperone network and the cytoskeleton. In summary, using a cell culture model for malignant transformation analyzed with 2D-PAGE, proteome and cellular changes can be related to defined steps of tumorigenesis.

show abstract

Section: Discussionmentioning

confidence: 99%

“…100 W until the running front reached the end of the glass plate. Gels were fluorescence stained with RuPBS as described elsewhere [23,24]. Scanned gels were compared and analyzed with the image analysis software PDQuest Advanced 8.0.1 (Bio-Rad, Munich, Germany) with BJ cells taken as control.…”

Section: Methodsmentioning

confidence: 99%

Malignant transformation in a defined genetic background: proteome changes displayed by 2D-PAGE

et al. 2010

Self Cite

View full text Add to dashboard Cite

show abstract

“…The band visualization can also be realized through an electrostatic binding of colored staining agents (such as Coomassie Brilliant Blue; CBB []) to certain residues (such as amino group) of proteins. Similarly, in the fluorometric method, there are fluorescent staining agents (such as SYPRO® Ruby [] and RuBPS []) that bind to protein by electrostatic interaction and there are also nonfluorescent or weakly fluorescent chemicals that form strong luminescent species when the chemicals react irreversibly with certain groups of the proteins. An example of the latter case is the bis‐cyclometalated solvento iridium(III) complexes [] that form strong luminophore after the histidine residues of proteins replace the labile solvento ligands.…”

Section: Methodsmentioning

confidence: 99%

“…Commercially available fluorescent dyes can be divided into two categories based on their chemical natures, that is, organic dyes (such as epicocconone [], Flamingo™ (http://www.bio-rad.com), and Krypton™ (http://www.thermoscientific.com)) and metal complex or metal chelate dyes (such as SYPRO® Ruby [] and RuBPS []). Metal chelates are chemically and photochemically more stable than organic dyes.…”

Section: Methodsmentioning

confidence: 99%

Luminescent iridium(III) complexes as novel protein staining agents

2012

View full text Add to dashboard Cite

This article reports a new class of luminescent metal complexes, biscyclometalated iridium(III) complexes with an ancillary bathophenanthroline disulfonate ligand, for staining protein bands that are separated by electrophoresis. The performances of these novel staining agents have been studied in comparison with tris(bathophenanthroline disulfonate) ruthenium(II) tetrasodium salt (i.e. RuBPS) using a commercially available imaging system. The staining agents showed different limits of detection, linear dynamic ranges, and protein-to-protein variations. The overall performances of all three stains were found to be better than or equivalent to RuBPS under the experimental conditions.

show abstract

“…In addition, not all fluorescent stains are compatible with the various gel types that are used to fractionate proteins for LC-MS/MS. Ruthenium II, which is much cheaper than SYPRO stains, causes increased background staining in Bis-Tris gels relative to Tris-Glycine gels (Moebius et al, 2007). These are all factors to keep in mind as part of your experimental design.…”

Section: Fractionation and Gel Stainingmentioning

confidence: 99%