A multiple high-resolution mini two-dimensional polyacrylamide gel electrophoresis system: Imaging two-dimensional gels using a cooled charge-coupled device after staining with silver or labeling with fluorophore

Urwin, Valerie E.; Jackson, Peter

doi:10.1016/0003-2697(91)90290-a

Cited by 28 publications

(9 citation statements)

References 19 publications

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“…Since labels such as monobromobimane require covalent modification of proteins, they are often less suitable for Edman-based protein sequencing and MALDI-TOF-mass spectrometry than their noncovalent counterparts [21,24]. When performing 2-D gel electrophoresis, prederivaElectrophoresis 2000, 21, 2509±2521 Luminescent protein stain for SDS-PAGE 2519 Figure 6.…”

Section: Discussionmentioning

confidence: 99%

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex

Berggren¹,

Chernokalskaya

Steinberg³

et al. 2000

Electrophoresis

337

205

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SYPRO Ruby dye is a permanent stain comprised of ruthenium as part of an organic complex that interacts noncovalently with proteins. SYPRO Ruby Protein Gel Stain provides a sensitive, gentle, fluorescence-based method for detecting proteins in one-dimensional and two-dimensional sodium dodecyl sulfate-polyacrylamide gels. Proteins are fixed, stained from 3h to overnight and then rinsed in deionized water or dilute methanol/acetic acid solution for 30 min. The stain can be visualized using a wide range of excitation sources commonly used in image analysis systems including a 302 nm UV-B transilluminator, 473 nm second harmonic generation (SHG) laser, 488 nm argon-ion laser, 532 nm yttrium-aluminum-garnet (YAG) laser, xenon arc lamp, blue fluorescent light bulb or blue light-emitting diode (LED). The sensitivity of SYPRO Ruby Protein Gel Stain is superior to colloidal Coomassie Brilliant Blue (CBB) stain or monobromobimane labeling and comparable with the highest sensitivity silver or zinc-imidazole staining procedures available. The linear dynamic range of SYPRO Ruby Protein Gel stain extends over three orders of magnitude, which is vastly superior to silver, zinc-imidazole, monobromobimane and CBB stain. The fluorescent stain does not contain superfluous chemicals (formaldehyde, glutaraldehyde, Tween-20) that frequently interfere with peptide identification in mass spectrometry. While peptide mass profiles are severely altered in protein samples prelabeled with monobromobimane, successful identification of proteins by peptide mass profiling using matrix-assisted laser desorption/ionization mass spectrometry was easily performed after protein detection with SYPRO Ruby Protein Gel stain.

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Section: Discussionmentioning

confidence: 99%

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex

Berggren¹,

Chernokalskaya

Steinberg³

et al. 2000

Electrophoresis

337

205

View full text Add to dashboard Cite

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“…In 2D electrophoresis, they must be grafted either between IEF and SDS-PAGE (Urwin and Jackson 1991) or after electrophoresis. In the latter case, molecules which are non-fluorescent but become fluorescent after coupling are generally chosen, such as fluorescamine (Jackowski and Liew 1980), or 2-methoxy-2,4-diphenyl furanone (MDPF) (Jackson et al 1988).…”

Section: Introductionmentioning

confidence: 99%

Ruthenium II tris (bathophenanthroline disulfonate), a powerful fluorescent stain for detection of proteins in gel with minimal interference in subsequent mass spectrometry analysis

et al. 2000

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Fluorescent ruthenium chelates have been prepared and investigated for staining proteins separated by electrophoresis. Ruthenium II tris (bathophenanthroline disulfonate) appears to be suitable for detection of proteins, with sensitivities in the nanogram range, although sensitivity does not reach the level of optimized silver staining methods. This compound can be efficiently detected either with UV tables or with commercial laser fluorescence scanners. A further advantage of this detection method is its compatibility with mass spectrometry., These factors make this method a good compromise for proteomics studies.

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“…The first and oldest mechanism is covalent binding, quite often of probes that are not fluorescent but become so when the covalent binding takes place (45,46). While the performances of such probes were not very impressive, and thus of limited use, a quantum leap was achieved when probes with much higher light absorption and emission characteristics were used.…”

Section: Protein Detection Via Fluorescencementioning

confidence: 99%

The Whereabouts of 2D Gels in Quantitative Proteomics

Rabilloud

2012

Methods in Molecular Biology

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Two dimensional gel electrophoresis has been instrumental in the development of proteomics. Although it is no longer the exclusive scheme used for proteomics, its unique features make it a still highly valuable tool, especially when multiple quantitative comparisons of samples must be made, and even for large samples series. However, quantitative proteomics using two-dimensional gels is critically dependent on the performances of the protein detection methods used after the electrophoretic separations. This chapter therefore examines critically the various detection methods, (radioactivity, dyes, fluorescence and silver) as well as the data analysis issues that must be taken into account when quantitative comparative analysis of two-dimensional gels is performed.

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A multiple high-resolution mini two-dimensional polyacrylamide gel electrophoresis system: Imaging two-dimensional gels using a cooled charge-coupled device after staining with silver or labeling with fluorophore

Cited by 28 publications

References 19 publications

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex

Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex

Ruthenium II tris (bathophenanthroline disulfonate), a powerful fluorescent stain for detection of proteins in gel with minimal interference in subsequent mass spectrometry analysis

The Whereabouts of 2D Gels in Quantitative Proteomics

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