TC FlAsH: Expression Analysis Detection Kits
Protein detection using TC-FlAsH Expression Analysis Detection Kits—Red (cat# A10068) 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 (cat# 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 contrasting stain in either red or orange allows visualization of your total protein alongside the Benchmark™ total protein ladder for reference.
- 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|