The obtained MC-BMP2 was solubilized in 50 μL of distilled water (DW). One microliter of NuPAGE LDS buffer (4×) (Invitrogen; CA, USA) and 2.5 μL of NuPAGE reducing agent (10×) (Invitrogen; CA, USA) were added to 6.5 μL of MC-BMP2 solution. For the control, 1 μL of NuPAGE LDS buffer and 2.5 μL of NuPAGE with 500 mM dithiothreitol as reducing agent were added to 2 μL of native BMP2-OpgY (1 μg/μL) solution. Native BMP2-OpgY and MC-BMP2 were individually incubated at 70 °C for 15 min and then loaded onto a NuPAGE 4–12% Bis-Tris gel (Invitrogen; CA, USA), followed by electrophoresis at 200 mV using XCell SureLock (Invitrogen; CA, USA). MC-BMP2 and native BMP2 were visualized by staining with 0.025% Coomassie Blue.
Nupage reducing agent 10
Manufactured by Thermo Fisher Scientific
Sourced in United States
The NuPAGE reducing agent (10×) is a concentrated solution designed to reduce disulfide bonds in proteins prior to electrophoresis. It is intended for use with the NuPAGE protein electrophoresis system.
Automatically generated - may contain errors
Lab products found in correlation
Xcell surelock, by Thermo Fisher Scientific (1 mentions)
Nupage 4 12 bis tris gel, by Thermo Fisher Scientific (1 mentions)
Nupage lds buffer, by Thermo Fisher Scientific (1 mentions)
Image lab software, by Bio-Rad (1 mentions)
Chemidoc touch imager, by Bio-Rad (1 mentions)
Mes buffer, by Thermo Fisher Scientific (1 mentions)
Simplyblue safestain, by Thermo Fisher Scientific (1 mentions)
Precision plus protein ladder, by Bio-Rad (1 mentions)
2 protocols using nupage reducing agent 10
1
SDS-PAGE Analysis of MC-BMP2
2
Protein Sample Preparation and Analysis
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Protein samples were prepared for analysis by adding water, NuPage Reducing Agent (10×) (Invitrogen, Carlsbad, CA), and NuPage LDS Buffer (4×) (Invitrogen) for a total of 10 μL per well. Non-reduced samples were prepared similarly, substituting 1 μL of water for the reducing agent. Each sample was put into a NuPage™ 4–12% Bis-Tris PAGE gel (Invitrogen) along with a PrecisionPlus™ protein ladder (Biorad, Hercules, CA). Gels were run in MES buffer (Invitrogen). After electrophoresis, gels were rinsed three times with 18 MΩ water for 5 min each. Gels were then treated with SimplyBlue™ Safestain (Invitrogen) for at least one hour, and then rinsed with water overnight. Images of stained gels were taken using a Chemidoc Touch Imager (BioRad). Densitometry measurements were performed using ImageLab software (BioRad). Densitometry readings of >96% were considered to be pure enough to proceed with animal trials.
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