Western Technical Support
NOTE: For the best possible results, Cell Signaling Technology (CST) strongly recommends using our optimized application-specific protocols for each product. These protocols are the result of extensive in-house validation performed at CST and ensure accurate and reproducible results.
To ensure a timely solution, please have the information below ready when you contact us. Technical Support can be reached by toll free phone at 877-678-8324 or 877-678-TECH, fax at 978-867-2400 or e-mail at email@example.com.
Have you consulted our Western Immunoblotting Troubleshooting Guide before using this technical report? For a listing of recommended cell types and treatments for use as positive controls for our activation state antibodies, visit our Recommended Controls page.
- Catalog number.
- Lot number and reference date printed on tube.
- How long have you had the product?
- At what temperature was the product stored?
- Was the product aliquotted?
- Cell type used for the sample lysate: (cell line, primary, whole tissue).
- Sample species.
- Induction/drug treatment used.
- Concentration of the drug used.
- What were the time points of induction?
- Positive control used.
- Lysis buffer used: (SDS Sample Buffer, CST Cell Lysis Buffer, CST Chaps Buffer, CST RIPA Buffer, other).
- What specific phosphatase inhibitors, if any, were included in the lysis buffer?
- Were the samples sonicated?
- Amount of protein lysate loaded per well.
- Date of lysate preparation.
- Lysate storage temperature: (4°C, -20°C, -80°C, other).
- Has this antibody previously worked on these samples?
- Percentage of acrylamide in the gel.
- Which membrane was used: (Nitrocellulose, PVDF, Nylon, other).
- What is the pore size of the membrane being used for transfer?
- Components of the blocking buffer.
- What was the blocking time?
Primary Antibody Incubation
- Primary antibody dilution.
- Components of the primary antibody dilution buffer.
- Incubation time for the primary antibody.
- Washing buffer and time after primary antibody incubation.
Secondary Antibody Incubation
- Secondary antibody source.
- Secondary antibody dilution.
- Components of the secondary antibody dilution buffer.
- Incubation time for the secondary antibody.
- Washing buffer and time after secondary antibody incubation.
- Which detection system was used: (Horseradish Peroxidase (HRP), Alkaline Phosphatase (AP), other).
- Supplier of the detection system.
- Length of time for film exposure.
- What is the appearance of the exposed and developed film?