Chromatography Forum

I'm testing some units and all of them seem to be giving me low signal. They are for A1c testing and the A10 peak is lower than required. The instrument uses ion affinity columns. Any help figuring out what is wrong would be greatly appreciated.

The flow rate in these units seems fine. They are within specifications and the sample loop fills completely. It is it is a straight shot to the column after that. Also I can't see any leaks.

I determined the blood is fine after testing it on a working unit in the lab.

I currently don't have any reason to suspect the detectors. With a flow through column they perform fine. Detector response also appears normal.

What I have found is that only certain columns have enough signal. The only issue with this is that I went through 8 brand new columns to have 7 of them not work. Also many other columns that have worked in the past aren't currently working. All signs are pointing to the columns being the issue. I took a column that worked and tested it on multiple instruments and got the same results on each instrument. The only problem is the incredibly small likelyhood that all these columns spontaneously went bad.

Does anyone have some suggestions of something I could try or comment on my diagnoses of the issue being columns?

Thanks in advance

Look at the efficiency/plate count. The column that performs satisfactory might be a bit more efficient than the others.
An optimization of the separation/column temperature might be a viable step to take. The mobile phase/s can be optimized as well – f. x. the ion strength or pH. Finally a hardware optimization (plumbing etc.) reducing excessive dead volume would be another important action.

Best Regards

Do you use a standard HPLC instrument or a dedicated diagnostic instrument?
Do you use dedicated columns from the instrument manufacturer or standard Affinity columns?
Together with the column you received a COA. Please try to reproduce the same test on your instrument as described on the COA. If you don't get the minimum similar result please contact the column manufacturer. It is not impossible that even after QC at the manufacturer due to for example high temperature all columns are effected.