Application of dyes in polyacrylamide upper gel electrophoresis and im-munoblotting
10.3969/j.issn.1000-4718.2025.05.023
- VernacularTitle:染料在聚丙烯酰胺上层凝胶电泳免疫印迹中的应用
- Author:
Weiguang CHEN
1
;
Chenjie YAN
;
Zhaojin XU
;
Yiting DONG
;
Zhichao MAO
;
Jiawei SUN
;
Wantie WANG
;
Linbo YUAN
Author Information
1. 温州医科大学组织胚胎学教研室,浙江 温州 325035
- Publication Type:Journal Article
- Keywords:
sodium dodecyl sulfate-polyacrylamide gel electrophoresis;
Remazol BrOrange yellow;
erythro-sine;
immunoblotting
- From:
Chinese Journal of Pathophysiology
2025;41(5):1036-1040
- CountryChina
- Language:Chinese
-
Abstract:
AIM:To investigate the effect of dyes,Remazol BrOrange yellow(RBY)and erythrosine(ERY),on the outcomes of immunoblotting analysis when used for staining the concentrate gel in sodium dodecyl sulfate-polyacryl-amide gel electrophoresis(SDS-PAGE).METHODS:Polyacrylamide gels were divided into five groups:the control group(prepared according to the conventional kit protocol),the RBY-stained group with a final concentration of 0.08 g/L,the RBY-stained group with a final concentration of 0.16 g/L,the ERY-stained group with a final concentration of 0.08 g/L,and the ERY-stained group with a final concentration of 0.8 g/L.Gels were prepared and subjected to electro-phoresis,followed by coomassie brilliant blue staining to visualize protein bands.Subsequently,proteins were transferred to PVDF membranes,which were then blocked,incubated with primary and secondary antibodies,washed,and finally ex-posed for imaging to observe the target protein vinculin bands.RESULTS:Compared with the unstained concentrate gel,the loading wells of the RBY or ERY pre-stained concentrate gel were more clearly visible.Analysis of the gels stained with coomassie brilliant blue after electrophoresis and marker visualization showed no significant different in protein elec-trophoretic mobility between prestained and unstained gels.Comparative analysis of the immunoblotting also indicated that the detection of protein samples transferred to PVDF membranes was unaffected.CONCLUSION:Prestaining concen-trate gels with RBY or ERY can enhance the efficiency of gel-based electrophoresis and immunoblotting analysis.
