Chromatography Forum

Hi all,
I had recently come accross a intresting problem which i could not able to solve yet.
I was analysing a protein compound(inclusion bodies). It is a basic analytical method using C4 column with water and Acetonitril TFA. When i injected the standard i got the response which i usually get but, for the sample i got only half the response as compared to my regular response. As a matter of fact, i had just injected in to another HPLC wherein, i got correct sample response as expected. I do not find any difference between two HPLC's(both are Agilent 1100). My sample is slightly viscous as compared to the standard so, the i first thing i checked was injector draw speed & eject speed but both the HPLC's were set same(100µl/min).

Can any one comment on this?

One system to another? Could be something other than autosampler. First question is "how is the precision of replicates" ?

Tubing dimensions, tubing dead volumes, detector cell differences, VWD v. DAD, binary v. quad pump, going through thermostatted compartment tubing, one having a column-switching valve, lots of things could be the reason.

Along with the thoughts presented by CPG - needle offset? Maybe one is at 1 mm and the other at 0, and you happened to bottom out in the 0-offset vial? Or perhaps you have a leaking metering seal, or had a partially plugged injection seat, or maybe a failing rotor seal? Like the man said, there can be a lot of factors. If you can consistently take a standard from one instrument to the other and obtain similar results, but taking a sample (I mean the exact same vial in both cases!) between the two results in a 50% difference, then this is a possibly more involved problem. Hopefully you re-prepped and re-injected your sample on both instruments?