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HIS Lite™ iFluor® 647 Tris NTA-Ni Complex
Example protocol
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles
Prepare a 5 to 10 mM stock solution by adding an appropriate amount of ddH2O.
Note: Store any unused stock solution at -20 °C. Avoid repeated freeze-thaw cycles and minimize light exposure.
PREPARATION OF WORKING SOLUTION
Prepare a 1 to 10 µM HIS Lite™ iFluor® 647 Tris NTA-Ni Complex working solution in PBS.
Note: Ensure that there is sufficient working solution to fully submerge the gel. After use, discard the working solution. Do not reuse.
SAMPLE EXPERIMENTAL PROTOCOL
The following protocol should be used only as a guideline and may require optimization to better suit your specific experimental needs.
Run gels based on your standard protocol.
Place the gel in a suitable container. Fix the gel in the fixing solution for 60 minutes. Note: 40% ethanol + 10% acetic acid can be used as a fixing solution.
Wash the gel twice with the ultra-pure water.
Incubate the gel in the HIS Lite™ iFluor® 647 Tris NTA-Ni Complex working solution for 60 minutes.
Note: Be sure to fully submerge the gel in the working solution.
Remove the working solution and wash the gel twice with PBS.
Proceed to imaging the gel immediately.
Mix the His-tagged protein solution and the HIS Lite™ iFluor® 647 Tris NTA-Ni Complex working solution at the appropriate concentrations.
Note: Optimization of the HIS Lite™ iFluor® 647 Tris NTA-Ni Complex to the His-tagged protein mix must be performed for better labeling.
Note: 1 to 10 µM of HIS Lite™ iFluor® 647 Tris NTA-Ni Complex can be used as a starting concentration.
Note: The reaction can be performed in a buffer containing 50 mM HEPES/KOH, pH 7.4, 100 mM KCl, 1 mM MgCl2, 2 mM β-mercaptoethanol, 5% glycerol, or a buffer of your choice.
Mix can be incubated for 30 minutes at room temperature or 4 ℃.
Note: Optimization of the incubation time and conditions must be performed for better labeling
Mix can then be subjected to column purification or any other downstream process.
Spectrum
Product family
| Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield | Correction Factor (260 nm) | Correction Factor (280 nm) |
| HIS Lite™ iFluor® 568 Tris NTA-Ni Complex | 568 | 587 | 1000001 | 0.571 | 0.34 | 0.15 |
References
Authors: Shafifar, Mina and Mozhgani, Sayed-Hamidreza and Razavi Pashabayg, Kobra and Mosavat, Arman and Karbalaei, Mohsen and Norouzi, Mehdi and Rezaee, Seyed Abdolrahim
Journal: Life sciences (2022): 120920
Authors: Crauwels, Maxine and Van Vaerenbergh, Nele and Kulaya, Neeme Benedict and Vincke, Cécile and D'Huyvetter, Matthias and Devoogdt, Nick and Muyldermans, Serge and Xavier, Catarina
Journal: New biotechnology (2020): 20-28
Authors: Raducanu, Vlad-Stefan and Isaioglou, Ioannis and Raducanu, Daniela-Violeta and Merzaban, Jasmeen S and Hamdan, Samir M
Journal: The Journal of biological chemistry (2020): 12214-12223
Authors: Pan, Xiaoshu and Yang, Yu and Li, Long and Li, Xiaowei and Li, Qiang and Cui, Cheng and Wang, Bang and Kuai, Hailan and Jiang, Jianhui and Tan, Weihong
Journal: Chemical science (2020): 9648-9654
Authors: Yang, Hui and Xiong, Haoran and Mi, Kaihang and Zhang, Yingying and Zhang, Xiaojun and Chen, Guohong
Journal: Fish & shellfish immunology (2020): 62-68
Safety data sheet