Western Blot Transfer Buffer, 10X, IC-7908
Product Introduction
This product is a 10X concentrated stock solution of Western blot transfer buffer, the standard buffer used for the electrophoretic transfer of proteins from polyacrylamide gels to membranes in Western blotting procedures. When diluted to 1X working concentration with deionized water and supplemented with the appropriate amount of methanol or ethanol, the buffer provides the ionic strength, pH buffering capacity, and conductivity necessary for efficient and quantitative electroblotting of proteins onto nitrocellulose or PVDF membranes. This Tris-Glycine-based transfer buffer is compatible with both tank (wet) and semi-dry transfer systems and is suitable for transferring proteins across a broad molecular weight range. The 10X concentrated formulation provides convenience for long-term storage and allows rapid preparation of working solutions, ensuring lot-to-lot consistency and reproducible transfer efficiency.
Product Features
1. Standard Western Blot Transfer Buffer: Optimized 10X formulation of Tris-Glycine transfer buffer for efficient protein electroblotting.
2. Ready-to-Dilute Concentrate: Supplied as a 10X stock requiring only dilution with deionized water and addition of methanol or ethanol for 1X working solution preparation.
3. High Transfer Efficiency: Provides consistent conductivity and buffering capacity for quantitative protein transfer onto nitrocellulose or PVDF membranes.
4. Broad Compatibility: Suitable for both tank (wet) transfer and semi-dry transfer systems and compatible with a wide range of protein molecular weights.
5. Convenient Storage: 10X concentrated format minimizes storage footprint and extends shelf life.
Specifications
Size: 500 mL
Formulation: 10X Concentrate (250 mM Tris, 1.92 M Glycine, pH 8.3-8.5)
Storage and Stability
Storage Conditions: Store at room temperature (15-30¡ãC).
Shelf Life: The product is stable for 24 months from the date of manufacture when stored as directed.
Protocol (For Reference Only)
Important: Freshly prepare 1X working transfer buffer with the appropriate alcohol concentration. Pre-chill transfer buffer to 4¡ãC for optimal transfer of high molecular weight proteins.
1. Preparation of 1X Working Transfer Buffer: Dilute the 10X Western Blot Transfer Buffer 1:10 with deionized water (e.g., 100 mL of 10X buffer plus 700 mL of deionized water), add 200 mL of methanol (for a final concentration of 20% methanol) or 150 mL of ethanol (for a final concentration of 15% ethanol), and mix thoroughly to obtain 1X transfer buffer.
2. Gel Equilibration: Following SDS-PAGE, carefully remove the gel from the glass plates and equilibrate the gel in 1X transfer buffer for 5-10 minutes at room temperature to remove electrophoresis buffer salts and SDS.
3. Membrane Preparation: Activate PVDF membrane by brief immersion in 100% methanol followed by equilibration in 1X transfer buffer for at least 5 minutes; for nitrocellulose membranes, equilibrate directly in 1X transfer buffer.
4. Transfer Assembly: Assemble the transfer sandwich in the following order: sponge or fiber pad, filter paper, gel, membrane, filter paper, sponge or fiber pad, ensuring complete removal of air bubbles between each layer. Place the sandwich in the transfer cassette with the membrane facing the anode.
5. Electrophoretic Transfer: Fill the transfer tank with ice-cold 1X transfer buffer, insert the cassette, and transfer at constant current of 200-350 mA for 1-2 hours at 4¡ãC with stirring, or at constant voltage of 100 V for 1 hour. For semi-dry transfer, assemble the sandwich and transfer at 10-15 V for 30-60 minutes depending on gel thickness and protein size.
6. Post-Transfer: After transfer is complete, disassemble the sandwich, rinse the membrane briefly in TBS or PBS, and proceed to blocking and immunodetection.
Precautions
1. Methanol is flammable and toxic; handle with appropriate safety precautions in a fume hood. Ethanol may be substituted at 15% final concentration.
2. Do not add methanol directly to the 10X concentrate; always dilute the 10X concentrate with water first, then add alcohol to prevent buffer component precipitation.
3. Pre-chill transfer buffer to 4¡ãC before use to dissipate heat generated during high-current transfer and prevent gel swelling or protein degradation.
4. Ensure complete removal of air bubbles from the transfer sandwich to prevent localized transfer failure ("bald spots") on the membrane.
5. For research use only. Not for use in diagnostic or therapeutic procedures.
FAQ (Simplified)
Q1: Why is methanol or ethanol added to the transfer buffer?
A1: Alcohol facilitates the removal of SDS from proteins and improves protein binding to the membrane. For high molecular weight proteins, reducing methanol to 10% may improve transfer efficiency; for low molecular weight proteins, maintaining methanol at 20% is recommended.
Q2: Can I reuse the 1X transfer buffer?
A2: 1X transfer buffer is typically used once for optimal transfer efficiency. Reuse is not recommended as alcohol evaporates and buffer ions deplete, reducing transfer efficiency and reproducibility.
Q3: What transfer conditions are recommended for different protein sizes?
A3: For proteins below 100 kDa, transfer at 100 V for 1 hour or 200-250 mA for 1.5-2 hours. For proteins above 100 kDa, transfer at lower current (100-200 mA) overnight at 4¡ãC to ensure complete transfer without overheating.
Q4: Can this buffer be used for semi-dry transfer?
A4: Yes. This 10X concentrate diluted to 1X with 20% methanol is fully compatible with semi-dry transfer systems. Adjust transfer time according to gel thickness and manufacturer recommendations.
Disclaimer
1. For Research Use Only. Not for use in diagnostic or therapeutic procedures.
2. Due to the variable nature of biological research, optimization of transfer conditions is recommended for specific protein targets and membrane types.
3. This warranty is limited to the replacement of the product. The manufacturer assumes no liability for incidental or consequential damages, including loss of samples or data.
4. Wear appropriate protective clothing and gloves when handling this product.
Ordering Information
Catalog Number: IC-7908
Product Name: Western Blot Transfer Buffer, 10X
Size: 500 mL
Price: CNY ¥200.00 / USD $20.00 / EUR €24.00 / JPY ¥3600.00
2023 Version
