K&A - Understanding Trityl Monitors

This article explains how the trityl monitor works, how to interpret its readings, and why it's a key tool in monitoring oligonucleotide synthesis efficiency.

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🔍 What is a Trityl Monitor?

The trityl monitor is a built-in photometer that helps you evaluate coupling quality during synthesis. It measures how much trityl cation (a bright orange byproduct of successful deblocking) is present in each column’s waste stream.

• Wavelength: Measurements are taken at 470 nm (blue light), where trityl absorbs strongly.

  

• Location: Each column is monitored independently in its own cuvette.

  

• Units: Output is not absolute OD—it's a relative extinction value ranging from ~1 (strong trityl signal) to ~4095 (no trityl detected).

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🔧 Key Concepts

V2 Reference Value

• V2 is the extinction value of the second trityl readout (cycle 2).

  

• All future values are compared to this to evaluate relative coupling strength. For That reason, on the histogram the V2 Value is always 100%

  

• A low V2 value (e.g., 10–50) = strong trityl signal = good early coupling.

• If the initial coupling is poor and the V2 Value is high, the V2 reference will be skewed, which can lead to misleading-looking charts even if the synthesis is failing.

Tip: Always validate your V2 range by comparing it to downstream QC results. A typical V2 range is 1–100, but this can vary depending on your chemistry and protocols.

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Total Yield

Used to track how much product remains after each synthesis cycle. It’s based on changes in extinction values compared to the baseline:

Formula:

 

• A higher extinction reading (closer to 4095) means less trityl retained → lower yield.

• If extinction values don’t change from cycle to cycle, yield stays at 100%.

• REMEMBER: This is in reference to the V2 value.  It is still recommended that you post-process and very yields downstream.

Example:

• Baseline = 1700, Cycle 20 = 3000 → Yield drops to 45.7%

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ASWY – Average Stepwise Yield

This tells you how efficient each coupling step is.

Formula:

ASWY (%)=(Total Yield)1/Number of Cycles X 100

 

Example:
45.7% total yield after 20 cycles → ASWY = 97.45% per step

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📄 Viewing Trityl Readings in Log Files

In addition to the monitor chart, you can find detailed trityl readings in the instrument log files, which are especially useful for troubleshooting or long-term tracking.

Each trityl reading entry typically includes:

• $C1 – $C6: Correspond to individual synthesis columns

• Baseline: The V2 reference set early in synthesis (cycle 2)

• Current: The latest extinction value recorded by the monitor

• Percent: Calculated yield for that column at this cycle

Example from Log: 

This means column 3 had a good initial coupling (baseline 19), but at the current cycle, the extinction value has increased to 614 — indicating trityl retention has dropped and the cumulative yield is ~85.4%.

Tip: Use these log values to identify coupling failures early, even if the synthesis completes. Large jumps in trityl extinction can point to running out of amidites, clogging, faulty valves, or chemistry issues.

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🧪 Protocol Setup Tips

• At the moment, the logs are not saved.  If you want to collect that data for future reference you can take screenshots

• The first deblock of each cycle is sent to the trityl monitor.

• To avoid measuring trityl from unnecessary steps (like secondary deblocks or washes), put those steps in a later section of your protocol.

• You can optionally add an extra deblock at the end of each cycle so that the main deblock (which carries the trityl) goes directly to waste instead of the monitor.

The trityl monitor is automatically purged at the end of every cycle. Purge duration is configured in the software. This value will typically be 20 seconds for most systems and 30-40seconds for VLS Systems.

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🖱️ Navigating the Chart

• Left click & drag down-right to zoom in

• Left click & drag up-left to zoom out

• Right click to scroll

• Each chart is independent—you can zoom and scroll separately for each column.

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✅ Summary