6-TAMRA (6-Carboxytetramethylrhodamine) serves as a popular fluorophore for the creation of bioconjugates, particularly in the synthesis of fluorescent antibodies and avidin derivatives employed in immunochemistry.

6-TAMRA labeled nucleotide experiment
Operation steps
1. Sample treatment: Incubate in NHOH/methylamine (1:1) solution at 65°C for 10 minutes to separate oligonucleotides from CPG.
2. Take the supernatant and wash CPG with 1 mL EtOH/MeCN/H2O (3:1:1).
3. Mix the supernatants and dry them.
4. Treat with fresh anhydrous triethylammonium fluoride/n-methylpyridone (250 μL, n-methylpyridone 1.5 mL, triethylamine 750 μL, tea-3hf 1.0 mL) at 65°C for 1.5 h to remove t-butyl-dimethylsilyl protecting groups from RNA residues, and add 25 μL 3 M NaOAc and 1 mL n-BuOH to precipitate oligonucleotides.
5. The sample was cooled at -70°C for 1 h and then centrifuged at 10,000 g for 30 min. The supernatant was decanted, washed with aqueous EtOH (70% v/v), and then dried.
6. 6-TAMRA (0.1 mL, 10 mg/mL in dimethyl sulfoxide) was added to the 3'-amino-modified oligonucleotide suspended in 1.0 mL of sodium bicarbonate buffer (pH 8.5) and incubated at 37°C for 12 h.
7. The labeled oligonucleotide was resuspended in water and passed through a G25 Nap-10 disposable desalting column to remove free dye.
8. The oligonucleotide was purified by HPLC with a linear gradient of acetonitrile in 0.1 M triethylammonium acetate (TEAA) buffer, pH 7.2.
9. The entire sample was loaded on a Hamilton PRP-1 column and eluted with a linear gradient of acetonitrile for 40 min. Samples were monitored at 260 and 297 nm, and peaks corresponding to the dual-labeled oligonucleotide species were collected.