Ab227074

Manufactured by Abcam

Sourced in United States

Ab227074 is a laboratory consumable product offered by Abcam. It is a single-use item designed for use in scientific research and experiments.

Automatically generated - may contain errors

Lab products found in correlation

7 protocols using ab227074

Western Blot Analysis of PPAR and IKZF Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

The MM cells were harvested, washed with phosphate-buffered saline (PBS), and resuspended in a lysis buffer containing 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 1% sodium deoxycholate, and 0.1% sodium dodecyl sulfate (SDS). The cells were lysed via brief sonication and then spun down to remove the cell debris. The total protein in the cell lysates was quantified using a Dc protein estimation kit (Bio-Rad, Hercules, CA, USA) with bovine serum albumin (BSA) as a standard. The cell lysates were loaded and run on SDS-polyacrylamide gel electrophoresis. The proteins were transferred onto a nitrocellulose membrane. The membrane was blocked with 5% BSA in tris-buffered saline containing 0.05% Tween 20 (TBST) and incubated with primary antibodies against PPARα (ab227074; Abcam, Waltham, MA, USA), PPARβ/δ (A5656; Abclonal, Woburn, MA, USA), PPARγ (16643-1-AP; Thermo Fisher, Waltham, MA, USA), IKZF1 (NBP1-98314; Novus Biologicals), and IKZF3 (NBP2-46048; Novus Biologicals, Littleton, CO, USA) in TBST containing 5% BSA overnight at 4 °C with gentle rocking. The membrane was then probed with an HRP-conjugated secondary antibody and developed using a Pierce ECL substrate.

Wu J., Wang X., Zhang M., Mathews P, & Kang Y. (2023). RXR Agonists Enhance Lenalidomide Anti-Myeloma Activity and T Cell Functions while Retaining Glucose-Lowering Effect. Cells, 12(15), 1993.

+ Open protocol

+ Expand

Protein Expression Analysis in MM Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

MM cells were harvested, washed with phosphate-buffered saline (PBS) and resuspended in a lysis buffer containing 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 1% sodium deoxycholate, and 0.1% sodium dodecyl sulfate (SDS). Cells were lysed via brief sonication. The lysates were centrifuged at high speed for 15 min to remove the cell debris. Total protein was quantified using a Dc protein estimation kit (Bio-Rad) with bovine serum albumin (BSA) as a standard. Approximately 20 μg of protein was loaded and subjected to SDS-polyacrylamide gel electrophoresis. The proteins were transferred onto a nitrocellulose membrane. The membrane was blocked with 5% BSA in tris-buffered saline containing 0.05% Tween 20 (TBST) and primary antibodies against PPARα (ab227074; Abcam), PPARβ/δ (A5656; ABclonal), PPARγ (16643-1-AP; Thermo Fisher), IKZF1 (NBP1-98314; Novus Biologicals), and IKZF3 (NBP2-46048; Novus Biologicals) were incubated with 5% BSA in TBST overnight at 4 °C with gentle rocking. The membrane was then probed with an HRP-conjugated secondary antibody and developed using a Pierce ECL substrate.

Matsushita K., Shinagawa E., Adachi O, & Ameyama M. (1990). Cytochrome a1 of acetobacter aceti is a cytochrome ba functioning as ubiquinol oxidase.. Proceedings of the National Academy of Sciences of the United States of America, 87(24), 9863-9867.

+ Open protocol

+ Expand

ChIP Assay Protocol for Transcription Factors

Check if the same lab product or an alternative is used in the 5 most similar protocols

ChIP assays were performed according to the manufacturer’s protocol (#91820; Cell Signaling Technology, Danvers, MA). Briefly, the cells were collected and cross-linked with 1% formaldehyde. After centrifugation, the resulting pellets were sonicated and the chromatin solution was precleared with 30 μL of CHiP-Grade protein G magnetic beads (#9006; Cell Signaling Technology). The soluble fraction was collected, and the chromatin beads were incubated with positive control histone H3 rabbit (#4620; Cell Signaling), normal rabbit IgG (#2729; Cell Signaling), anti-PPARα (ab227074; Abcam), anti-PPARγ (16643-1-AP; Proteintech Rosemont, IL), anti-FLAG M2 (F1804; Sigma; for PPARβ/δ with a FLAG tag) antibodies at 4 °C overnight. ChIP-enriched DNA was analyzed by quantitative PCR using the CRBN promoter primers as follows: forward: 5′-TCCCCTGTCACCTTCAAAAC-3′ and reverse: 5′-TGCCTTGTGAGTCTGACACC-3′. The enrichment of specific genomic regions was assessed relative to the input DNA, followed by normalization to the respective control IgG values. Percent input = 4% × 2^{(C[T] 4% input sample-C[T] IP sample)}.

+ Open protocol

+ Expand

Protein Expression Analysis in Placental Tissues

Check if the same lab product or an alternative is used in the 5 most similar protocols

Sixteen placental tissues (control groups, n = 8; ICP groups, n = 8) were lysed with RIPA lysis buffer (ASPEN) in the presence of protease inhibitor, and then, the quantification of the isolated protein was quantified and evaluated utilizing using the BCA Protein Assay Kit (Pierce, Rockford, IL, USA). The proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride membranes. Non-specific binding was blocked with 5% skimmed milk for 1 h at room temperature. Next, the membranes were incubated with anti-APOA2 (1:1000, Abcam, ab92478); anti-PPARα (1:1000, Abcam, ab227074) as the primary antibody at 4°C overnight, respectively, followed by a secondary antibody (1:5000, Affinity, Shanghai, China) at room temperature for 30 min. Beta-actin (1:2000, Abcam, ab8227) was used as an internal control. The antigens and antibodies were detected using high-signal ECL substrates, and the gray values of protein bands were analyzed and quantified using Image J.

Zeng W., Hou Y., Gu W, & Chen Z. (2024). Proteomic Biomarkers of Intrahepatic Cholestasis of Pregnancy. Reproductive Sciences, 31(6), 1573-1585.

+ Open protocol

+ Expand

Protein Profiling via RIPA Lysis and Western Blot

Check if the same lab product or an alternative is used in the 5 most similar protocols

Radioimmunoprecipitation assay (RIPA) solution encompassing protease inhibitors (Solarbio) was used to lysate total cell proteins. The proteins were then transferred to polyvinylidene difluoride (PVDF) membranes (Roche, Basel, Switzerland) after being separated on an SDS–polyacrylamide gel. At room temperature, the membranes were blocked with 5% fat-free dry milk before being cultivated with primary antibodies with diluted 2000 times. The membranes were then treated with the secondary antibody for 2 h at room temperature. ECL-enhanced chemiluminescence reagents (Amersham, Little Chalfont, UK) were used to visualize protein bands. Primary antibodies used in this investigation included anti-PDK4 (ab110336; Abcam, Cambridge, UK), anti-MMP-3 (ab52915; Abcam), anti-MMP-13 (ab39012; Abcam), anti-ADAMTS-4 (ab314856; Abcam), anti-PPARA (ab227074; Abcam), anti-PPARD (ab178866; Abcam), anti-ACSL1 (ab177958; Abcam), and anti-GAPDH (ab52915; Abcam).

Li Z., Xie L., Zeng H, & Wu Y. (2024). PDK4 inhibits osteoarthritis progression by activating the PPAR pathway. Journal of Orthopaedic Surgery and Research, 19, 109.

+ Open protocol

+ Expand

Isolation and Staining of Cardiomyocytes

Check if the same lab product or an alternative is used in the 5 most similar protocols

Harvested hearts were placed in a Langendorff setup and perfused with a Type II Collagenase and Protease digestion buffer and stepped up with Calcium to 1 mM. Viable CMs were sorted using LP-FACS^{12 (link)}. The Union Biometrica COPAS Flow Platform was used to select CMs based on extinction, time of flight, and red fluorescence. For staining, hearts were flash frozen into O.C.T Compound blocks (Fisher Healthcare 23-730-571) and stored at −80 C. Samples were sliced into 10 um sections using a cryo-microtome and placed on glass slides. Primary antibodies against 1:500 dilution α-actinin (Abcam ab227074) were used along with 1:1000 dilution DAPI and 1:200 dilution Wheat Germ Agglutinin counterstains. A Leica SP8 confocal microscope running Leica Application Suite X was used to image slides with a ×63 oil immersion objective.

Murphy S.A., Miyamoto M., Kervadec A., Kannan S., Tampakakis E., Kambhampati S., Lin B.L., Paek S., Andersen P., Lee D.I., Zhu R., An S.S., Kass D.A., Uosaki H., Colas A.R, & Kwon C. (2021). PGC1/PPAR drive cardiomyocyte maturation at single cell level via YAP1 and SF3B2. Nature Communications, 12, 1648.

+ Open protocol

+ Expand

Protein Expression Analysis in Membranes

Check if the same lab product or an alternative is used in the 5 most similar protocols

Detailed descriptions are given as previously described [25] . Briefly, the membranes were blotted with antibodies against PPARα (Abcam, Cambridge, MA, USA, #ab227074), GAPDH (Abcam, #ab181602), and FABP4 (Abcam, #ab 92501).

Sun C., Song B., Sheng W., Yu D., Yang T., Geng F., Fang K., Jiao Y., Zhang J, & Zhang S. (2022). Fenofibrate Attenuates Radiation-Induced Oxidative Damage to the Skin through Fatty Acid Binding Protein 4 (FABP4). Frontiers in bioscience (Landmark edition), 27(7).

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!