Carryover is mostly caused by: poor HPLC maintenance (a lack of), sticky samples or poor solubility. Simple things such as making sure your sample is completely soluble in the mobile phase. Seems straightforward enough, but soooo many people ignore this item so it really is the number one problem I see with clients.

Another item which can effect carry over is the type of Autoinjector used. The design of the autoinjector itself can have a major impact on carryover as some are more prone to having problems than others. The best designs utilize a true flow-through wash system where the mobile phase if continuously flushed through the A/I flushing away all residues. Example: Most of newer Agilent's autoinjectors (i.e. 1100/1200/1260/1290) use a high pressure flow-through design which washes the ENTIRE injector's active flow path ALL of the time (during the method). The needle, need seat, loop, high pressure sampler pump and valve are continuously flushed throughout the method. No wash vials or rinsing of the injector are needed for 99.9% of samples. IMHO: The best design out there and I am not affiliated with Agilent at all, I just appreciate great design and engineering. Most every other manufacturer uses either a low pressure injector system (i.e. syringe) or partially flushed (i.e. Waters) design which requires the use of wash vials and solutions to remove any residues. *I rarely am called in to fix carryover issues with flow through designs and when I am, it is almost always due to a worn out Autoinjector rotor seal (lack of maintenance) or sample precipitation (not the fault of the instrument).

I have attached a link to a free article that I wrote on this topic, "Carry-Over (Carryover) Contamination in HPLC and LC-MS Systems", which may help you.

http://hplctips.blogspot.com/2015/02/carry-over-carryover-contamination-in.html

Hello!

The basic problem of carryover of proteins in LC-MS is also the column. Alone with gradient, Solvents, temperature, and pressure you will not solve this problem. Follow the advice of Bill Letter! In addition, however, you should see if you have the correct column. Also a C4 column can not always avoid the problem of carryover. You might consider using a monolithic column or a column with very large pore size, as a major problem of carryover may also be the fibrils of the proteins.

Best regards

Burghard Cordes

Hi!

You can try a PEEK needle versus a  steel needle. Also changing the composition of the strong wash and the weak wash may help.

Best Regards,

Lucia Neuman