- Rapid, direct labeling of purified or crude cell lysate means no gel staining needed
- Visualize your protein of interest in gel and even within the cassette alongside reference Benchmark ™ protein ladder as well as total protein in contrasting color
- Westerns are no longer necessary to confirm TC-tagged proteins
Our TC-FlAsH Expression Analysis Detection Kits are a rapid and easy way to confirm the presence of your protein of interest tagged with a tetracysteine (TC) tag. The simple setup permits the use of crude or purified cell lysate, requires 13 minutes of prep time to stain the proteins for loading, and better still, permits visualization of your protein of interest in gel and even within the cassette, and no more gel handling or gel staining mess. The contrasting stain in either red or orange allows visualization of your total protein alongside the Benchmark™ total protein ladder for reference.
Protein detection using TC-FlAsH Expression Analysis Detection Kits in polyacrylamide gel. Images are of TC-tagged proteins, both purified and in crude lysate, and Benchmark ladder run on a SDS-PAGE 4-20% Tris-glycine gel. Green pseudocolor is for TC-tagged protein detection, obtained with 473nm excitation with 520nm long pass emission filter and red pseudocolor is for total protein detection, obtained with 633nm excitation with 675nm long pass emission filter on a Fuji FLA3000 laser scanner. With FlAsH detection, only TC-tagged proteins appear with the Benchmark Fluorescent Protein Ladder (LC5928). With total protein detection, all proteins present are detected, including TC-tagged proteins. No post-staining was performed and gels were able to be imaged in-cassette.
- The small 6-amino acid 1 kD tetracysteine tag (CCPGCC) and FlAsH stain described by Roger Tsien offer the high specificity, high sensitivity, and flexibility needed for protein labeling.
- Molecular Probes® now offers tools for rapidly identifying TC-tagged proteins in gels as well as in-cell for microscopy.
- Imaging will require a Safe Imager, UV box, or laser scanner and appropriate filters.
Table 1 - The specificity of the TC-Tag provides accuracy as well as sensitivity
|TC-FlAsH vs. Other Tags||TC-FlAsH||Covalys SNAP tag||LUMIO||Invision His-tag|
|Sensitivity in cassette||at 488: 4 ng
at UV: 60 ng
|Sensitivity out of cassette||at 488: 250 pg
at UV: 1 ng
|at 488: < 20 pg||at UV: 30 ng||at UV: 5 ng|
|Total protein sensitivity at either 665 or 620 nm||Orange: IN 100 ng/OUT 12 ng
Red: IN 100 ng/ OUT 50 ng
|Detection||Excitation peak (nm)||Emission peak (nm)||Filter recommendation|
|TC-tagged protein||500 mid-UV||535 mid-UV||520LP
|Orange total protein||580 mid-UV||620 mid-UV||580LP
|Red total protein||650 mid-UV||665 mid-UV||675LP|