One of the most sensitive enhanced chemiluminescent substrates available.
Thermo Scientific SuperSignal West Femto Substrate is an ultra-sensitive enhanced chemiluminescent (ECL) substrate for low-femtogram-level detection by Western blot analysis.
SuperSignal West Femto Substrate Maximum Sensitivity Substrate is an ultra-sensitive chemiluminescent substrate system for Western blotting with horseradish peroxidase (HRP) enzyme. The substrate kit enables detection of low femtogram (that's mid-zeptomole!) amounts of protein deposited on nitrocellulose or PVDF membrane and probed with appropriate primary and secondary antibodies. When combined with optimized antibody concentrations and blocking buffers, SuperSignal West Femto Substrate enables detection of target proteins in amounts that are small to be seen with typical ECL substrates.
ECL substrate – an enhanced chemiluminescent substrate for detection of horseradish peroxidase (HRP) activity from antibodies and other Western blot probes
Sensitive – detect low-femtogram (mid-zeptomole) amounts of protein in bands on nitrocellulose or PVDF membranes
Quantitative – produces quantitative signal that is measurable over two orders of magnitude
Saves on antibody – optimized for very dilute antibody concentrations; dilute primary antibodies 5000- to 100,000-fold and secondary antibodies 100,000 to 500,000-fold (from 1mg/mL stocks)
Intense signal – easy to capture an image by exposure to autoradiography film or via a CCD camera or similar imaging system
Excellent signal duration – 8 hours of usable light-output when conditions are optimized
Stable reagent – kit components are stable for 1 year at 4°C or 6 months at room temperature
True femtogram levels of detection with Thermo Scientific SuperSignal West Femto Maximum Sensitivity Substrate. Purified IκB was serially diluted from 100 to 1fg and then electrophoresed on a 4-20% mini gel. The protein was transferred to PVDF membrane and blocked with Thermo Scientific StartingBlock Blocking Buffer for 1 hour at room temperature. The blot was incubated in rabbit anti-IκBα (1mg/mL) at 1:1000 dilution overnight at 4°C, followed by incubation in HRP-conjugated goat anti-Rabbit IgG (1mg/mL) at 1:200,000 dilution for 1 hour at room temperature. The membrane was exposed to Thermo Scientific CL-XPosure Film for 1 minute.
Use less antibody with Thermo Scientific SuperSignal West Femto Maximum Sensitivity Substrate. MAP Kinase and HSP86 were detected in HeLa cell lysate prepared by dilution in Laemmli sample buffer and heating to 95°C for 5 minutes. Serial dilutions (1:1) of total protein (10µg to 19.5ng) were separated by electrophoresis and the proteins transferred to nitrocellulose membranes (Part No. 88013). Thermo Scientific Starting Block in TBS Blocking Buffer (Part No. 37542) was used for blocking. Rabbit anti-MAP Kinase (Millipore Part No. 06-182) and Rabbit anti-HSP86 (Part No. PA3-013) were used as primary antibodies at 0.2µg/mL (1:5000 dilution of 1mg/mL stock solution). Goat anti-Rabbit HRP (Part No. 31460) was used as secondary antibody at 2ng/mL (1:500,000 dilution of 1mg/mL stock solution). MAP Kinase and HSP86 targets were detected using Thermo Scientific SuperSignal West Femto (Part No. 34095) and Thermo Scientific CL-XPosure Film (Part No. 34091).
Thermo Scientific SuperSignal West Femto is more sensitive than GE Healthcare ECL Plus Substrate. Two-fold serial dilutions of mouse IL-2 were detected with both substrates. The primary antibody dilution was 1/2000 and the secondary antibody dilution was 1/300,000 (from 1mg/mL stocks).