5X Protein Loading Buffer (Non-Reducing)

Product Identifier

Introduction

5X Protein Loading Buffer (Non-Reducing) is very convenient to use in SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and Western blotting. This protein loading buffer contains a bright pink tracking dye instead of Bromophenol blue. This loading buffer is supplied as a 5X stock and should be diluted before application.

Description

5X Protein Loading Buffer (Non-Reducing) is a unique loading buffer due to the presence of a pink tracking dye which runs as the dye front during SDS-PAGE and transfers completely to the nitrocellulose membrane while performing Western blotting protocol. This visible pink band helps to determine the efficiency of transfer as well as to align the cut membrane. The 5X concentration of this buffer facilitates the loading of larger sample volume per well compared to the traditional 2X loading buffer. This loading buffer contains SDS which is required for disruption of all higher order protein structures and glycerol for proper loading of the protein sample.

Application

5X Protein Loading Buffer (Non-Reducing) can be used as a loading buffer in all kinds of denaturing gels and are compatible with coomassie and silver staining and Western blotting. The marker dye transfers and binds only to nitrocellulose. The dye does not transfer and bind to nylon or PVDF membranes. The pink dye auto fluoresces and, therefore, might interfere with fluorescence detection applications.

5X Protein Loading Buffer (Non-Reducing) is very convenient easy to use. One part of loading buffer has to be mixed with four parts of protein sample before loading. The entire electrophoretic procedure can be monitored through tracking the pink dye front.

Procedure

1. Allow the 5X Protein Loading Buffer (Non-Reducing) to come to room temperature.
2. Mix one volume of Loading Buffer with four volumes of protein sample.
3. Boil the sample for 3-5 minutes.
4. Quick chill on ice. Centrifuge at 10,000 RPM for one minute.
5. Load the sample into the gel and perform the electrophoresis.
6. Stop electrophoresis when the pink tracking dye reaches the bottom of gel.

Composition

5X Protein Loading Buffer (Non-Reducing) is composed of 0.3M Tris HCl, 5% SDS, 50% glycerol, and a proprietary pink tracking dye.

Properties

Storage conditions

The 5X Protein Loading Buffer (Non-Reducing) has to be stored at 15-25 °C. Under recommended condition, the reagent is stable for 12 months.

Warning and Precautions

Not for Medicinal Use. Read the SDS carefully before beginning the protocol. Wear protective gloves/protective clothing/eye protection/face protection. Follow good clinical laboratory practices while handling clinical samples. Standard precautions should be followed as per established guidelines. Safety guidelines may be referred in safety data sheets of the product.

Performance and Evaluation

Performance of the solution is expected when the solution is stored at recommended temperature and within the expiry period.

Safety Information

The 5X Protein Loading Buffer (Non-Reducing) is for laboratory use only, not for drug, household or other uses. Take appropriate laboratory safety measures and wear gloves and safety goggles when handling. Not compatible with disinfecting agents containing bleach. Please refer the Safety Data Sheet (SDS) for information regarding hazards and safe handling practices.

Technical Assistance

At HiMedia we pride ourselves on the quality and availability of our technical support. For any kind of technical assistance, mail at mb@himedialabs.com.