Use of Activators


Applicable Products

0234 BTT (Crystalline)
0237 ETT (Crystalline)
3140/2 ETT (0.25M)
3145/6 ETT (0.5M)
3160/2 BTT (0.3M)

Traditionally, tetrazole was used as the activator during oligonucleotide synthesis. Although this was chemically efficient for DNA synthesis, there were several drawbacks. Tetrazole has a tendency to precipitate from aceonitrile solution due to its low solubility. This is especially true in winter months, where it was not uncommon for tetrazole to precipitate on the synthesiser overnight resulting in blocked lines. Coupling efficiency during RNA synthesis, or while using other sterically hindered phosphoramidites, was greatly reduced in comparison to DNA synthesis. Finally, tetrazole is classed as an explosive which makes transportation of the product, even in solution, costly or impossible.

There are many alternative activators available today, the most common used being 0.25M ETT (3140/3142), 0.3M BTT (3160/3162) and DCI (3150). All of these are much more soluble than tetrazole in acetonitrile thus resolving the crystallisation problem. In terms of coupling efficiency in DNA synthesis, they are all comparable with tetrazole. However, they are much more efficient when used with sterically hindered phosphoramidites such as 2’-OTBDMS RNA monomers. In fact, using 0.3M BTT has been reported to reduce the coupling time of 2’-OTBDMS protected RNA amidites to 3min rather than the 12-15min required when using tetrazole. In RNA synthesis, using BTT as an activator has brought coupling efficiencies in line with DNA synthesis, i.e >99%. The same applies to the use of 0.5M ETT (3145/3146).

One drawback is using ETT or BTT rather than tetrazole is that they are more acidic (pKa: ETT 4.3; BTT 4.1; tetrazole 4.89). This can be problematic when synthesising on a larger scale (>10-15μmol) or for very long oligos. In this case DCI (pKa 5.2) can be used to avoid depurination or loss of trityl groups during coupling (please note that we no longer offer DCI).

Preparation of BTT & ETT Solutions

Rather than use pre-mixed solutions, some customers prefer to prepare their own activator solutions. This gives the flexibility of making only the quantity required for synthesis. There is also the added advantage that shipping the solids does not incur a hazardous charge.

Crystalline BTT (0234) and ETT (0237) are provided in three pack sizes (1g, 10g and 25g) for dilution to customers’ required concentrations in whatever synthesiser bottle is appropriate. Typical dilution information is given below (these examples equate to routine bottle sizes and concentrations, however, obviously a range of pack sizes is provided to give customers flexibility in this).


Mol. Wt.



BTT (0.3M) 192.24 11.53g 25.95g
ETT (0.25M) 130.17 6.51g 14.64g
ETT (0.5M) 130.17 13.02g 29.29g

To prepare the solution simply weigh out the desired amount of product into your instrument bottle and dilute by adding filtered, anhydrous acetonitrile (Diluent, 4050) as indicated. Ensure that the solid is entirely dissolved prior to use. This is essential to avoid blockages on the synthesiser. Use end line filters on the activator bottle position, again to prevent any blockages.

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