School of Pharmacy UNIVERSITY OF WISCONSIN - MADISON Sample Preparation Guidelines Do’s DO keep sample amounts low. For most applications on our LC-MSD VL instrument we can detect a few hundred picograms, so concentrations in the low tens of micrograms per microliter are usually sufficient. If you are working with a new compound on the APCI and don’t know how it will behave, start with a very low concentration and go higher until you see a signal. DO make the sample as clean as possible. Purity and amount are the two big determinants for getting good mass spectrometry results. DO work with volatile solvents and buffers. Volatile buffers will pass through the mass spectrometer. Non-volatile buffers, salts etc., leave residue in the source, in the capillary and in the mass spectrometer necessitating costly, time-consuming clean outs. DO ask questions. Gary and Cameron are happy to consult with all the users regarding any and all aspects of mass spectrometry. We are glad to help with designing experiments etc. if it means cleaner samples and higher quality data. It is always better to ask than risk damaging columns or instruments by using inappropriate techniques or reagents. DO centrifuge your samples before loading them in the vials for mass spectrometric analysis. Particulates clog columns leading to high back pressures. Filtration is another option but centrifugation is preferred. Don’t’s DO NOT operate the APCI instrument (or others) without appropriate training. Training for the walk-up APCI LC-MSD and other instruments is offered frequently throughout the year. If you require mass spectrometry for your project and you have not been trained, ask another member of your lab or another lab to run the samples for you until you complete the training. DO NOT use trifluoroacetic acid (TFA). While TFA is an excellent pairing reagent and gives nice peak shape for HPLC, it dramatically reduces ionization of most compounds at the source. DO NOT try to analyze samples with high concentrations of mineral salts (Na, K etc.). Excess non-volatile buffer components can be deposited on the source during ionization and reduce sensitivity. They can also build up in other parts of the instrument and cause problems. DO NOT analyze samples with high concentrations of organic solvents. You could damage or dissolve the matrix, ruining the column. DO NOT analyze samples with corrosive compounds in them. Perchloric acid and other corrosives will damage or destroy the stainless steel parts of the HPLC system.