Anti akr1c3

Manufactured by Merck Group

Sourced in United States

Anti-AKR1C3 is a laboratory reagent used for the detection and quantification of the AKR1C3 protein in biological samples. AKR1C3 is an enzyme involved in various metabolic processes. This product can be used in research applications that require the analysis of AKR1C3 expression or activity.

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Lab products found in correlation

Anti pparγ, by Merck Group (1 mentions) Anti human gapdh, by Merck Group (1 mentions) Gel doc analysis system, by Bio-Rad (1 mentions) β actin, by Merck Group (1 mentions) Dc protein assay, by Bio-Rad (1 mentions) Vectastain elite abc kit, by Vector Laboratories (1 mentions) Diaminobenzidine substrate kit, by Vector Laboratories (1 mentions) Bx51 microscope, by Olympus (1 mentions) Ecl chemiluminescent substrate, by Cytiva (1 mentions) Cycloheximide (chx), by Selleck Chemicals (1 mentions) Anti bcl 2 sc 7382, by Santa Cruz Biotechnology (1 mentions) Mg132, by Selleck Chemicals (1 mentions) Anti β actin, by Merck Group (1 mentions)

4 protocols using anti akr1c3

Western Blot Protein Expression Analysis

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Cultured cells were washed once with PBS buffer and then lysed in lysis buffer for 5 min. The sample was placed on ice for 30 min, and then centrifuged at 12,000 ×g for 30 min. We used the Bio-Rad DC protein assay to determine the protein concentration of each sample. Equal amounts of protein were separated by sodium salt-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidene fluoride (PVDF) membranes. We washed the PVDF membranes in Tween and Tris-Buffered Saline (TTBS) (20 mM Tris, 0.5 M NaCl, 0.05% Tween-20) for 10 min, and then blocked the membranes in 10% skim milk for 1 h. The primary antibodies were diluted with TTBS as follows: anti-AKR1C3 (1:4000, A6229–200U; Sigma-Aldrich Co.), anti-PPARγ (1:1000; Sigma-Aldrich Co.), β-actin (1:10,000), and anti-human GAPDH (1:3000). We used the Gel Doc analysis system (Bio-Rad) to scan the gray level of each band. Each Western blot was repeated at least twice.

Li X., Hong X., Gao X., Gu X., Xiong W., Zhao J., Yu H., Cui M., Xie M., Bai Y, & Sun S. (2018). Methyl jasmonate enhances the radiation sensitivity of esophageal carcinoma cells by inhibiting the 11-ketoprostaglandin reductase activity of AKR1C3. Cancer Management and Research, 10, 3149-3158.

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Immunohistochemical Analysis of Tumor Markers

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Tumors were fixed by formalin and paraffin embedded tissue blocks were dewaxed, rehydrated, and blocked for endogenous peroxidase activity. Antigen retrieving was performed in sodium citrate buffer (0.01 mol/L, pH 6.0) in a microwave oven at 1,000 W for 3 min and then at 100 W for 20 min. Nonspecific antibody binding was blocked by incubating with 10% fetal bovine serum in PBS for 30 min at room temperature. Slides were then incubated with anti-Ki-67 (at 1:500; NeoMarker), anti-AKR1C3 (at 1:100; Sigma) at 4°C overnight. Slides were then washed and incubated with biotin-conjugated secondary antibodies for 30 min, followed by incubation with avidin DH-biotinylated horseradish peroxidase complex for 30 min (Vectastain ABC Elite Kit, Vector Laboratories). The sections were developed with the diaminobenzidine substrate kit (Vector Laboratories) and counterstained with hematoxylin. Nuclear staining cells was scored and counted in 5 different vision areas. Images were taken with an Olympus BX51 microscope equipped with DP72 camera.

Liu C., Lou W., Zhu Y., Yang J.C., Nadiminty N., Gaikwad N.W., Evans C.P, & Gao A.C. (2015). Intracrine androgens and AKR1C3 activation confer resistance to enzalutamide in prostate cancer. Cancer research, 75(7), 1413-1422.

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Protein Expression Analysis by Western Blot

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Protein expression was compared by Western blot using mouse polyclonal anti-AKR1C3 (Sigma Co.). Briefly, cells were seeded in 6-well plates and cultured to 70% confluence. After the indicated treatments, cell protein extracts were prepared. Western blots were performed with 40 μg of protein extract and protein was detected using ECL chemiluminescent substrate (Amersham Pharmacia Biotech, Piscataway, NJ). GAPDH protein was used as a loading control. Western blot experiments were repeated at least three times.

Zhao J., Xiang Y., Xiao C., Guo P., Wang D., Liu Y, & Shen Y. (2014). AKR1C3 Overexpression Mediates Methotrexate Resistance in Choriocarcinoma Cells. International Journal of Medical Sciences, 11(11), 1089-1097.

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Protein Degradation and Ubiquitination Assay

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MG132 and cycloheximide (CHX) were purchased from Selleck Chemicals (Houston, TX, USA). All antibodies used in this study are listed below: Mouse monoclonal Anti‐β‐actin (A1978, Sigma‐Aldrich, St. Louis, MO, USA), anti‐AKR1C3 (A6229, Sigma‐Aldrich), anti‐AR‐V7 (AG10008, Precision Antibody, Columbia, MD, USA), anti‐ubiquitin (sc‐47721, Santa Cruz, Santa Cruz, CA, USA), anti‐Bcl‐2 (sc‐7382, Santa Cruz); rabbit polyclonal anti‐AR (N‐20) (sc‐816, Santa Cruz) and anti‐B4GALT1 (A8546, ABclonal, Woburn, MA, USA).

Wang B., Wu S., Fang Y., Sun G., He D., Hsieh J., Wang X., Zeng H, & Wu K. (2020). The AKR1C3/AR‐V7 complex maintains CRPC tumour growth by repressing B4GALT1 expression. Journal of Cellular and Molecular Medicine, 24(20), 12032-12043.

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