Protein Expression Analysis in Kidney Tissue

Kidney tissues were homogenized using radio-immunoprecipitation assay buffer (Wako Pure Chemicals, Osaka, Japan). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting were performed as described previously [24 (link)]. The primary antibodies against 5α-Red1 (Abcam, Cambridge, UK), 11β-HSD2 (Proteintech, Rosemont, USA), and β-actin (Abcam, located in Cambridge, UK) were used. The immunoreactive bands were visualized using Immobilon Western Chemiluminescent HRP Substrate (Millipore, Billerica, MA, USA). The intensities of the specific chemiluminescent bands were analyzed using an ImageQuant LAS 3000 (GE Healthcare, Amersham, UK).

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Nouchi Y., Munetsuna E., Yamada H., Yamazaki M., Ando Y., Mizuno G., Ikeya M., Kageyama I., Wakasugi T., Teshigawara A., Hattori Y., Tsuboi Y., Ishikawa H., Suzuki K, & Ohashi K. (2023). Maternal High-Fructose Corn Syrup Intake Impairs Corticosterone Clearance by Reducing Renal 11β-Hsd2 Activity via miR-27a-Mediated Mechanism in Rat Offspring. Nutrients, 15(9), 2122.

Publication 2023

radio-immunoprecipitation assay buffer β-actin Immobilon Western Chemiluminescent HRP Substrate ImageQuant LAS 3000

Actin Antibodies Assay Buffer Immobilon Immunoprecipitation Kidney Sds page Tissues

Corresponding Organization : Fujita Health University Hospital

Other organizations : Kagawa Prefectural College of Health Sciences, Tokyo University of Technology

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Variable analysis

independent variables

Tissue homogenization using radio-immunoprecipitation assay buffer

dependent variables

Protein expression levels of 5α-Red1, 11β-HSD2, and β-actin measured by Western blotting

control variables

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting protocols as described previously [24]
Primary antibodies used: 5α-Red1 (Abcam, Cambridge, UK), 11β-HSD2 (Proteintech, Rosemont, USA), and β-actin (Abcam, located in Cambridge, UK)
Visualization of immunoreactive bands using Immobilon Western Chemiluminescent HRP Substrate (Millipore, Billerica, MA, USA)
Analysis of chemiluminescent band intensities using an ImageQuant LAS 3000 (GE Healthcare, Amersham, UK)

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