Bovine pancreatic RNase A (Sigma Aldrich) was dissolved in 20 mM phosphate, 8 M urea, pH 8, to a concentration of ∼750 μM and then treated with DTT at a final concentration of 30 mM. The reaction mixture was incubated for 4 h at room temperature before quenching the reaction by lowering the pH to 3 with acetic acid (AcOH). The buffer was then exchanged to 0.1 M AcOH using PD10 columns. The protein concentration was determined and samples were aliquoted before lyophilization to dryness.
Bovine pancreatic rnase a
Manufactured by Merck Group
Sourced in United States, Germany
Bovine pancreatic RNase A is a laboratory enzyme derived from bovine pancreas. It is a ribonuclease that catalyzes the cleavage of RNA molecules.
Automatically generated - may contain errors
Lab products found in correlation
Propidium iodide, by Merck Group (2 mentions)
Live dead viability cytotoxicity kit, by Thermo Fisher Scientific (2 mentions)
Rnasealert kit, by Integrated DNA Technologies (2 mentions)
Bca protein assay kit, by BioTeke (2 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Modfit 3, by BD (1 mentions)
Annexin 5 fitc pi apoptosis detection kit, by Vazyme (1 mentions)
Bovine serum albumin (bsa), by Avantor (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Avantor (1 mentions)
Branched pei25k, by Merck Group (1 mentions)
Fluorescein isothiocyanate (fitc), by Merck Group (1 mentions)
Zinc acetate, by Merck Group (1 mentions)
2 methylimidazole, by Merck Group (1 mentions)
One step tunel cell apoptosis detection kit green fluorescence, by Beyotime (1 mentions)
Lactate dehydrogenase ldh cytotoxicity assay kit, by Beyotime (1 mentions)
Acetone, by Fujifilm (1 mentions)
Modfit software, by Verity Software House (1 mentions)
Human insulin, by Merck Group (1 mentions)
Bovine α la, by Merck Group (1 mentions)
Facscalibur cytometer, by BD (1 mentions)
8 protocols using bovine pancreatic rnase a
1
Bovine Pancreatic RNase A Preparation
2
miR-518b Impacts Cell Cycle and Apoptosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The role of miR-518b in cell cycle progression and apoptosis was detected by flow cytometry. According to the manufacturer's protocol, cells were cultured for 48 h, harvested and rinsed with phosphate-buffered saline (PBS), before being fixed in 70% ethanol at 4°C overnight. Subsequently, cells were rinsed with PBS again, and were treated with PBS containing 100 mg/l bovine pancreatic RNase A (Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) at 37°C for 2 h, followed by incubation with propidium iodide (Sigma-Aldrich; Merck KGaA) for 30 min at room temperature. Cell apoptosis was detected using the Annexin V fluorescein isothiocyanate Apoptosis kit (Calbiochem; EMD Millipore, Billerica, MA, USA) according to the manufacturer's protocol. Cell cycle progression and apoptosis were analyzed on a BD FACSCalibur flow cytometer with ModFit 3.0 software (both from BD Biosciences, Franklin Lakes, NJ, USA).
3
Cytotoxicity and Apoptosis Evaluation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Branched PEI25K (impurities: ≤1% water) and bovine pancreatic RNase A (≥70 kU/mg protein) were purchased from Sigma-Aldrich (St. Louis, MS, USA). Genipin (>98%) was provided by Zhixin Biotechnol. Co. (Linchuan, China). RNaseAlert® kit was obtained from Integrated DNA Technologies, Inc. (Coralville, IA, USA). FBS and DMEM were purchased from Kangyuan Co. (Beijing, China) and Gibco (Grand Island, NE, USA), respectively. BCA protein assay kit was provided by BioTeke Co. (Beijing, China). Blue plus II protein marker was purchased from TransGen Biotech. (Beijing, China). BSA and MTT were purchased from Amresco (Solon, OH, USA). LIVE/DEAD® Viability/Cytotoxicity kit and one-step TUNEL cell apoptosis detection kit were obtained by Thermo Fisher (Grand Island, NE, USA) and Beyotime (Jiangsu, China), respectively. The Annexin V-FITC/PI apoptosis detection kit was provided by Vazyme Co. (Nanjing, China).
4
RNase A Cytotoxicity Assays
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bovine pancreatic RNase A, zinc acetate, 2-methylimidazole, and fluorescein isothiocyanate (FITC) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Enzymatic activity was detected by RNase Alert® kit, which was obtained from Integrated DNA Technologies, Inc. (Coralville, IA, USA). BCA protein assay kit was obtained from BioTeke (Beijing, China). The LIVE/DEAD® Viability/Cytotoxicity kit was provided by ThermoFisher (Grand Island, NE, USA). One-step TUNEL cell apoptosis detection kit (green fluorescence) and lactate dehydrogenase (LDH) cytotoxicity assay kit were purchased from Beyotime (Jiangsu, China). Fetal bovine serum (FBS) and Dulbecco’s modified Eagle’s medium (DMEM) were purchased from Kangyuan Co. (Beijing, China) and Gibco (Grand Island, NE, USA), respectively. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 4′,6-diamidino-2-phenylindole (DAPI) were purchased from Amersco (Solon, OH, USA). All other reagents were of the highest grade available and used as received.
5
Preparation and Characterization of Bovine Pancreatic RNase A
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bovine pancreatic RNase
A, cCMP, CHCA, SA, and ANS were purchased from Sigma-Aldrich (St.
Louis, USA). Cholesterol (Wako special grade), CDI (for organic synthesis),
DMSO (guaranteed reagent), dehydrated DMSO (for organic synthesis),
DMSO-d6 (for NMR), and acetone (guaranteed
reagent) were purchased from FUJIFILM Wako Pure Chemical Industries,
Ltd. (Osaka, Japan). β-CyD was purchased from Nihon Shokuhin
Kako Co., Ltd. (Tokyo, Japan). All other reagents were commercial
products of analytical grade. Phosphate buffer (0.05 mol/L, pH 6.5)
was prepared by Na2HPO4 and KH2PO4.
A, cCMP, CHCA, SA, and ANS were purchased from Sigma-Aldrich (St.
Louis, USA). Cholesterol (Wako special grade), CDI (for organic synthesis),
DMSO (guaranteed reagent), dehydrated DMSO (for organic synthesis),
DMSO-d6 (for NMR), and acetone (guaranteed
reagent) were purchased from FUJIFILM Wako Pure Chemical Industries,
Ltd. (Osaka, Japan). β-CyD was purchased from Nihon Shokuhin
Kako Co., Ltd. (Tokyo, Japan). All other reagents were commercial
products of analytical grade. Phosphate buffer (0.05 mol/L, pH 6.5)
was prepared by Na2HPO4 and KH2PO4.
6
Cell Cycle Analysis of A549 and H1299 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A total of 1×106 A549 or H1299 cells were harvested, washed with ice-cold PBS and fixed in 70% ice-cold ethanol at 4°C overnight. The fixed cells were washed with PBS and resuspended in 1 ml PBS supplemented with 100 µg/ml bovine pancreatic RNase A (Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) and 40 µg/ml propidium iodide (PI; Sigma-Aldrich; Merck KGaA) for 30 min at 4°C, cell cycle analysis was performed with a Becton Dickinson FACSCalibur cytometer (BD Biosciences, Inc., Franklin Lakes, NJ, USA). Cell cycle analysis was performed using ModFit software (version 3.2.1, Verity Software House, Topsham, ME, USA).
7
Protein Purification Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bovine αLA, bovine pancreatic RNaseA, and human insulin were purchased from Sigma-Aldrich Co., LLC. (St. Louis, USA). Human ubiquitin was purchased from Funakoshi Co., Ltd. (Tokyo, Japan). Chemically synthesized human Aβ40 and human IAPP were purchased from Peptide Institute, Inc. (Osaka, Japan). All other reagents, including HEWL, were obtained from Nacalai Tesque, Inc. (Kyoto, Japan).
8
Cell Cycle and Apoptosis Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell cycle progression and cell apoptosis were performed using a Becton Dickinson FACS/Calibur cytometer (Becton Dickinson Biosciences, Inc., NJ, USA) after 48 h of incubation. For cell cycle analysis, the GBM cells were fixed in 70% cold ethanol followed by incubation with 2 μg/ml bovine pancreatic RNase A (Sigma, St. Louis, MO, USA) and 20 μg/ml propidium iodide (Sigma). Cell apoptosis was detected using the Annexin-V fluorescein isothiocyanate (FITC) Apoptosis Kit (Calbiochem, San Diego, CA, USA) according to the manufacturer’s protocol.
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?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!