3 - UPLC_Work_Practical_Considerations

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Running U-HPLC: Practical Considerations Injection Air Gap • Automatic or manual • Full loop or partial loop injection – Critical for partial loop injection Good Repeatability: Setting Up the Sample Manager • Full loop – Air gap • 4 µL for 30 µL needle • 2 µL for 15 µL needle – Overfill factor depends on loop size • see Help for Automatic parameters – Metering device compatible with sample volume • Partial loop – Low draw speed for low volume – Pre and post air gap are essential – Automatic parameters • Pre and post air gap of 1 µL • 100 µL per minute Shula Levin, Waters Israel 40
Running U-HPLC: Practical Considerations Strong Contaminations • If the system is contaminated, some spare parts must be changed – Why: • Due to previous bad or not appropriate washing, contamination has been accumulated and then released since the system is not able to compensate anymore • Manual washing is not strong enough to eliminate carry-over – How: Which spare parts must be changed • Seal wash of the wash station • Needle • VDD (bubble detector) • Loop Main Parameters Summary • Strong Washing – Strong solvent 200 µL • Weak Wash – Weak solvent 600 µL • Full loop or partial loop injection – Use automatic settings • Air gap – For partial injection – Automatic • Draw speed – Automatic Shula Levin, Waters Israel 41
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