Evaluation of GAPDH and β-actin as internal reference proteins of Skp2 by Western blot
10.13699/j.cnki.1001-6821.2024.06.013
- VernacularTitle:用蛋白质印迹评价GAPDH与β-actin作为Skp2内参蛋白的研究
- Author:
Jun-Yuan SUN
1
;
Wei-Wei LUO
;
Jin-Feng YANG
;
Jing JIA
Author Information
1. 杭州医学院检验医学与生物工程学院,浙江杭州 310013
- Keywords:
internal reference protein;
β-actin;
glyceraldehyde-3-phosphate dehydrogenase;
S-phase kinase-associated protein 2;
relative expression level;
Western Blot
- From:
The Chinese Journal of Clinical Pharmacology
2024;40(6):844-848
- CountryChina
- Language:Chinese
-
Abstract:
Objective To determine the appropriate internal reference protein for the Western Blot(WB)assay to detect proteins related to S-phase kinase-associated protein 2(Skp2).Methods This study was carried out with several human tumor cell lines.The intracellular Skp2 protein level was down-regulated by transient transfection of shRNA plasmid,and glyceraldehyde-3-phosphate dehydrogenase(GAPDH)and β-actin were used as internal reference proteins respectively for WB assay.The relative expression levels and correlations of GAPDH,β-actin and Skp2 were evaluated.Results When the relative expression level of Skp2 changed,the relative expression levels of GAPDH in human non-small cell lung cancer cells H1299(1.32 × 106±7.86 × 104),human cervical cancer cell HeLa(6.59 × 106±1.79 × 106)and human liver cancer cell HepG2(3.13 × 106±2.87 × 105)all changed significantly(all P＜0.05).The correlation results showed that for human non-small cell lung cancer cells A549(R=0.825 8)and H1299(R=0.906 2),and there was a certain correlation between Skp2 and GAPDH expression levels(all P＜0.05).Conclusion β-actin is more suitable to be applied as reference protein in WB assay for Skp2 protein.
