How long should needle and sample loop characterisation take?

<p>The reason I ask is that today I characterised after installation of a new 1µL loop and it took in excess of 25 minutes. Usually when I've tested with 2 and 5µL loops it is much shorter than this. Is there any underlying reasons why this might have taken so long this time around?</p>


  • Hello, I believe there are two reasons:

    Firstly, times do vary depending on the loop and needle installed. The characterization process must be done correctly to ensure accurate injection volumes, thus Sample Manager runs a slow syringe speed (10 µL/min) and checks the volume of the loop by locating the position of aspirated air bubble twice, once with the loop offline and once with the loop online. The 1 µL loop is a very small loop and the ID is smaller than the other loops, (larger loops are manufactured using 0.009 stainless steel) this means to transport the solvent through the loop is more difficult and the process is slower, like sucking on a small straw. The volume that the air gap needs to travel for a 30 µL needle and to the volume detection device is about 40 µL, so that is 4 minutes, or 8 minutes for the repeats. Additionally, the same tolerance issue, mentioned earlier becomes more pronounced as we make the loop's tubing ID smaller.

    The second reason is that if the Sample Manager gets an unsatisfactory answer with its first attempt it repeats, so I wonder if the process may have repeated. This uncertainty of detection of the aspirated air gap can arise if the mobile phase is UV absorbing, if contains TFA for example.

    I have data on times for the complete preparation of the Sample Manager at my fingertips, rather than characterization alone. At solvent changeover when one must perform:

    • Prime wash syringes (weak and strong)
    • Prime sample syringe
    • Characterize loop and needle volume

    The time for the default 30?L needle & 10?L loop is about 18 minutes and the time for the default 30?L needle & 2?L loop about 21 minutes.

  • TIM

    hi,in the document ,you mentioned the solvent go though the loop .whether the solvent is wash solvent or mobile phase

    whether i need perform the needle and sample loop characterisation if i chang the wash solvent or mobile phase

    thank you for your reply

  • lizh


    Yes, everytime the mobile phase changes you should recharacterize. The Sample Manager uses the needlewash (Weak) to pull through solvent and an air bubble to determine the loop size, so it is importnat to do regularly, particularly if using partial loop modes, PLPA and PLNO. Usually, most people set the "Sys Prep" function in the Console to do this automatically as part of the priming step and run it first thing.

    If you did not characterize the loop, the injection will be reproducible based upon the volume determined, but if your new wash solvent is radically different in viscocity, than using the existing calibration the system will pull a little less. It will do this perfectly reproducibly, but you will get less. The other reason to do the characterization is to determine if there is a leak.

    In short, once at the start of a work day or a new set of samples, but if you forget I would not be concerned, do so the next day and you will be fine. It must be done whenever a loop or needle is changed or when you did fix a leak.

    I hope that helps.


  • TIM

    thank you very much! lz