The TrueBlack® WB Blocking Buffer Kit is a ready-to-use buffer system for fluorescence-based western blotting (WB). The buffers yield optimal specificity and sensitivity by blocking non-specific interactions of dye-labeled antibodies with proteins and the blotting membrane.

TrueBlack® WB Blocking Buffer Kit Features

Blocks as well or better than Odyssey® Blocking Buffer, at a lower price
Reduces non-specific protein bands and background over entire membrane
Suppresses background from charged dyes better than BSA, gelatin, or casein
Compatible with PVDF and nitrocellulose membranes
Contains no mammalian proteins, for broad antibody compatibility
For visible and near-IR fluorescent westerns

Figure 1. Western detection of phospho-Erk1/2 in PDGF-stimulated NIH-3T3 cell lysate. Membranes were blocked with fish gelatin blocking buffer, LI-COR® Odyssey® TBS Blocking Buffer, or TrueBlack® WB Blocking Buffer. Phosphorylated Erk was detected using rabbit anti-pErk1/2 and CF®680R donkey anti-rabbit antibodies. TrueBlack® WB Blocking Buffer gave lower background fluorescence and better specificity compared to the other buffers.

Superior western blocking for next-generation fluorescent dyes

Non-specific signal in WB can arise from multiple sources, including antibody cross-reactivity with off-target proteins, non-specific antibody adsorption to the membrane, and membrane autofluorescence. Another potential cause of background is the effect of fluorescent dyes themselves on the specificity of labeled antibodies. Highly charged dyes like Alexa Fluor® or CF® dyes have improved solubility and brightness of conjugates compared to uncharged dyes. However, the extra charge carried by antibodies labeled with these dyes can result in non-specific binding to proteins and membranes. The TrueBlack® WB Blocking Buffer Kit blocks background from multiple sources including charged dye conjugates (Figure 2). TrueBlack® blocking buffer is especially advantageous for phosphoprotein detection, significantly improving specificity compared to conventional blocking buffers (Figure 1).

Switch from Odyssey® Blocking Buffer and save

TrueBlack® WB Blocking Buffer performs as well or better for fluorescent WB compared to LI-COR’s Odyssey® Blocking Buffer (Figure 1), and is priced lower on a per membrane basis.

Compare TrueBlack® WB Blocking Buffer Kit with Odyssey® Blocking Buffer

Product	TrueBlack® WB Blocking Buffer Kit	Odyssey® Blocking Buffer
Trial Size	For 10 membranes	125 mL for 4 membranes
Full Size	For 50 membranes	500 mL for 16 membranes

Number of membranes if Odyssey® Blocking Buffer is used for each blocking and antibody dilution step. TrueBlack® WB Blocking Buffer Kit includes enough buffers for all blocking and antibody incubation steps for the stated number of membranes.

Figure 2. Western detection of tubulin in HeLa cell lysate with mouse anti-tubulin and Alexa Fluor® 790 goat anti-mouse antibodies. Membranes were blocked with fish gelatin blocking buffer or TrueBlack® WB Blocking Buffer. The highly negatively charged Alexa Fluor® 790 labeled antibody showed non-specific binding to the PVDF membrane and cellular proteins, which was blocked by TrueBlack® WB Blocking Buffer. Lanes 1-3: 10 ug, 1 ug, or 0.1 ug HeLa cell total protein.Note: Alexa Fluor ®790 dye was used to demonstrate non-specific background from highly charged dyes. Biotium’s near-IR CF® dyes are peggylated, a structural feature that renders the dyes highly water soluble without excessive negative charge, resulting in significantly lower background.