SuperSignal™ Western Blot Substrate Trial Pack, Atto & Pico PLUS

Here are some suggestions:
- Shorten the time of exposure
- Wash blot and try using SuperSignal West Pico PLUS Chemiluminescent Substrate
- Strip the membrane using Restore Western Blot Stripping Buffer, re-probe with lesser amount of primary and secondary antibodies, and try first with SuperSignal West Pico PLUS Chemiluminescent Substrate
- Re-run the gel using lesser sample

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

Here are potential causes for high background and corresponding solutions:
- Too much HRP in the system: Further dilute the HRP-conjugate, see guidelines in Table 1 in the user guide (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0018858_SuperSignalWestAtto_UG.pdf).
- Inadequate blocking: Optimize blocking conditions.
- Inappropriate blocking reagent: Try a different blocking reagent. Use clear milk or StartingBlock blocking buffer instead of dry milk.
- Inadequate washing: Increase length, number or volume of washes.
- Film has been overexposed: Decrease exposure time or use Pierce Background Eliminator (Cat No. 21065).
- Concentration of antigen or antibody is too high: Decrease amount of antigen or antibody.
- Poor antibody specificity: Use SuperSignal Western Blot Enhancer (Cat. No. 46640).
We also recommend minimizing handling of the membrane, using clean gloves while handling, clean plasticware, glassware, forceps, or other contact surfaces. We also recommend trimming of the membrane to remove excess area.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

SuperSignal West Atto Ultimate Sensitivity Substrate is compatible with Restore Western Blot Stripping Buffer.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

We recommend using SuperSignal West Atto Ultimate Sensitivity Substrate for detection of proteins in the following situations:
- For proteins with low concentration, not measurable by BCA or Bradford assays
- For low-abundance phosphorylated or glycosylated proteins
- For low-abundance membrane or nuclear proteins
- For low-abundance proteins being detected with a low-affinity antibody or when the antibody is precious
- When sample source is limited or when multiple conditions or assays are being tested on a single sample

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

SuperSignal West Atto Ultimate Sensitivity Substrate can tolerate up to 3 freeze/thaw cycles without significant loss in performance.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.