How to choose the appropriate dwell time/cycle time in an MRM and sMRM method to get enough points per chromatographic peak.

For research use only. Not for use in diagnostic procedures.

Answer

Step 1: Make a first acquisition in MRM with a default cycle time (e.g. cycle time = 0.5 s)
Step 2: Determine the average width of the smallest chromatographic peak (for example 15 s at the base of the chromatographic peak)
Step 3: Calculate the cycle time with the following formula respecting the desired number of points per peak:
Cycle time = Peak width / Number of points per peak (e.g. cycle time = 15/10 = 1.5 s for 10 points/peak)

Step 4-a classical MRM: Determine the dwell time to be applied for each of the transitions in order to obtain the previously calculated cycle time (e.g. cycle time = 1.5 s).

For a fixed dwell time for each transition:
Cycle time = number of transitions x (Dwell time + Pause time between two MRM transitions)

• Dwell time = (Cycle time / Number of transition) - Pause time between two MRM transitions

Specifications for minimum dwell and pause times depend on the MS. Per default, the pause time is always set to 5 ms. (e.g. Dwell time = (1500/100)-5 = 10 ms for cycle time = 1.5s, pause time = 5ms and number of transition = 100)

For a variable dwell time:
Use the reference dwell time obtained with the previous calculation and increase/decrease the dwell time for the desired transitions. This decrease will have an impact on the other transitions because the cycle time for variable dwell times is calculated with the formula below:
Cycle time = Number of transition x Pause time between two MRM transitions + ∑ Dwell time for each transition

The following rule must be applied :

If the dwell time of "Y" MRM transitions is increased by "Z" ms then Dwell time of other transitions = reference dwell time - (Y x Z) / (Number of transition - Y)

Step 4-b in scheduled MRM: In sMRM the dwell time for each transition will vary according to the retention time because the number of transitions monitored at a given retention time will determine the dwell time. Analyst or SCIEX OS automatically calculates the correct dwell time for each MRM transition. It will take the largest dwell time value for each MRM transition that allows the desired cycle time determined in step 3 to be respected. Indeed, the higher the dwell time, the higher the signal to noise ratio. 

• The first parameter you must indicate is the desired cycle time which must respect the rules of step 3.

The size of the time windows is an important factor to consider for sMRM methods because the higher the number of transitions with the same retention time (RT ± ΔRT), the lower the dwell time used by the software. 

• The second parameter that you must indicate is the size of the time windows which must not be too large in order to limit the superposition of the MRM transitions, but which must not be too small to avoid the lack of robustness of the method (Retention time shift).
• The last parameter is to ensure that when creating an sMRM method the minimum dwell time for all MRM transitions is not less than the minimum dwell time of the MS. To check this, You must use the sMRM calculator script (Analyst) or sMRM Summary (SCIEX OS). If this criterion is not respected, it is necessary to reduce the size of the detection windows or/and to remove MRM transitions from the method.