Detection of Proteins in Polyacrylamide Gels Using Eriochrome Black T and Rhodamine B

Jung, Da Woon; Tak, Gyoung Hoon; Lim, Jin Wook; Bae, Chan Jin; Kim, Gil Young; Yoo, Gyurng Soo; Choi, Jung Kap

doi:10.1006/abio.1998.2813

Cited by 12 publications

(1 citation statement)

References 7 publications

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“…Alternate densitometric stains have not gained popularity within the proteomic research community because these stains have been unable to compete with the performance of fluorescent staining and imaging. Also, most of these alternate densitometric stains have, at best, had limited characterization or have similar detection sensitivity to cCBB and hence have not been pursued for further application in proteomics [54,[117][118][119][120][121][122][123][124][125][126].…”

Section: Additional Densitometric Stainsmentioning

confidence: 99%

Quantitative proteomics: assessing the spectrum of in-gel protein detection methods

2010

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Proteomics research relies heavily on visualization methods for detection of proteins separated by polyacrylamide gel electrophoresis. Commonly used staining approaches involve colorimetric dyes such as Coomassie Brilliant Blue, fluorescent dyes including Sypro Ruby, newly developed reactive fluorophores, as well as a plethora of others. The most desired characteristic in selecting one stain over another is sensitivity, but this is far from the only important parameter. This review evaluates protein detection methods in terms of their quantitative attributes, including limit of detection (i.e., sensitivity), linear dynamic range, inter-protein variability, capacity for spot detection after 2D gel electrophoresis, and compatibility with subsequent mass spectrometric analyses. Unfortunately, many of these quantitative criteria are not routinely or consistently addressed by most of the studies published to date. We would urge more rigorous routine characterization of stains and detection methodologies as a critical approach to systematically improving these critically important tools for quantitative proteomics. In addition, substantial improvements in detection technology, particularly over the last decade or so, emphasize the need to consider renewed characterization of existing stains; the quantitative stains we need, or at least the chemistries required for their future development, may well already exist.

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Section: Additional Densitometric Stainsmentioning

confidence: 99%

Quantitative proteomics: assessing the spectrum of in-gel protein detection methods

2010

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Counterion-dye staining method for DNA in agarose gels using crystal violet and methyl orange

Yang¹,

Jung²,

Bai

et al. 2001

Electrophoresis

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Sensitive and safe methods for visualization of DNA in agarose gels are described. 0.001% crystal violet dissolved in distilled water was used for DNA staining on agarose gels and it could detect as little as 16 ng of DNA (3 kb, pGem-7Zf/EcoRI) without destaining procedure. The detection limit is four times lower than that of ethidium bromide. To improve the sensitivity, we studied a counterion-dye staining method using methyl orange as a counterion-dye which contributes to reduce excessive background staining by crystal violet. Dye concentration, pH of staining solution, mixing molar ratio of two dyes, and staining times were optimized for the counterion-dye staining. By the staining with a mixed solution of 0.0025% crystal violet and 0.0005% methyl orange in distilled water, 8 ng of the 3 kb DNA in an agarose gel was detected within 30 min.

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Usefulness of visible dyes for the staining of protein or DNA in electrophoresis

Choi

2004

Electrophoresis

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Since 1993, we have studied visible organic dye stains for protein or DNA to improve methodologies and developed the counterion dye staining method. The method employs two oppositely charged dyes that form an ion-pair complex in the staining solution. The selective binding of free dye to protein or DNA in the staining solution improves detection sensitivity and speed. It is a rapid and sensitive procedure, involving fixing/staining or staining/quick destaining steps that are completed in 1-1.5 h. The lowest detection limits achieved are 4-8 ng of protein on polyacrylamide gels and approximately 10 ng of DNA on agarose gels. The focus of this review is to chronicle the development and current status of the counterion dye staining method for detection of protein or DNA. As an extended application of visible dyes, we also discuss the visible dye staining method for detecting protein on blotting membranes developed in our laboratory.

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Detection of Proteins in Polyacrylamide Gels Using Eriochrome Black T and Rhodamine B

Cited by 12 publications

References 7 publications

Quantitative proteomics: assessing the spectrum of in-gel protein detection methods

Quantitative proteomics: assessing the spectrum of in-gel protein detection methods

Counterion-dye staining method for DNA in agarose gels using crystal violet and methyl orange

Usefulness of visible dyes for the staining of protein or DNA in electrophoresis

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