Chromatography Forum

Hi,

I am currently working on a BIOQ triple quad MS to look for stress hormones and am not getting a good enough quanitication limit (need about 20%) better detection. I was wondering if MS/MS could provide an improvement and if so how much?

Also the machine I am working on is about 15 years old, would a newer machine give better detection and sensitivity?

Hope someone can give me some pointers as my PhD is resting on detecting these hormones!

Amy

A newer instrument will give you much better sensitivity.

How are you using your instrument at the moment? SIM?

Hi,

I am using SIM and searching for the four masses I need. Currently getting a quantification limit of about 0.4 ng/mL and need it to be 0.06! So frustrating! We don't have any newer machine in our lab and the MS/MS systems aren't working!

on the quad, if you are doing SIM, remember that the ion will loose energy along the way (traveling through the first quad, the collision cell and the third quad to the detector).
try and see if there is a setting for ion energy. that allways a good place to start fiddling in a quad. it might just give the ion that little boost it needs to get to the detector. also even if you are not fragmenting in the collision cell i have found that you need a little bit of collision energy to help the ion through to the third quad.
but MS/MS will definately increase your sensitivity

on the age of the instrument. if it was well maintained thoughout hte years and the quads was cleaned regularly you souldn't have a problem.
do you know what type of multiplier the instrument has. Electron multipliers loose their sensitivity sooner than a photomultiplier (or at least that what i've been told)

The actual signal in MS/MS may not increase, but because you are increasing selectivity, the noise should decrease. Since signal:noise is very often the limiting feature, it's definitely worth trying. You're not far from the target sensitivity! Make sure your instrument is properly calibrated too; if it has an electron multiplier then over the course of time this will lose some sensitivity, for which the instrument will probably compensate by gradually increasing the EM voltage, if you let it do its gain calibration.
Good luck.

Hi Amy, the BIOQ triple quad MS should already be an MS/MS instrument or does MS/MS not work on this machine? Since this is an old Micromass MS it should already have a photomultiplier detector so from that point sensitivity should be ok .
MS/MS will definitely increase sensitivity in a complex sample matrice since it will decrease noise. It should improve your LOD probably at least to a factor of 10.
The more up-to-date triplequad machines from Waters/Micromass all have the Z-Spray ionsource which also increases sensitivity but also robustness in dirty samples.
If you don't get your machine running to the performance you need you could look for another lab in your university which have a newer triple quad MS which you could possibly use for a limited time period. All the triples which are about 5 years old or younger should be from a factor of 50 to more than 100 more sensitive than your old BioQ and developping a MRM method for stress hormons should not take much time. Once you have got the method the analysis of the samples should really go fast...

Hi,

Thanks for your responses they are really helpful. I do indeed have a photomultiplier in the BIOQ and it is reassuring that this shouldn't have dropped sensitivity too much. I think we are trying to get the MS/MS working on the BIOQ (it previosuly wasn't) so hopefully we might be able to try this. It is really frustrating that I can't get the sensitivity on the method and we don't have a working MS/MS at the moment. Fingers crossed we will have one soon. I am wondering whether it is worth trying to go outside the uni to try the samples on a newer machine?

You cannot really say how much of a sensitivity increase you will get. However, I use a TQD regularly and seem to get ppb levels with MS SIM and ppt (trillion)-ppb with MS/MS. What happens is that you actual area counts will drop but the signal to noise ratio then jumps by a massive amount. So your actual instrument sensitivity will improve with that. That is if you correctly set the MS/MS experiments up properly. Remember that you should use unique collision energies per transition. So MS/MS is definitely worth it.

That allways a good place to start fiddling in a quad. it might just give the ion that little boost it needs to get to the detector.
Also even if you are not fragmenting in the collision cell i have found that you need a little bit of collision energy to help the ion through to the third quad.

Thanks...

Update: I am still struggling to get the detection limits I need, I am trying derivatisation on the MS, but we still don't have a working MS/MS

Also we don't have any newer machines here that I can try my method on which is very frustrustrating as looking at the literature it does seem that newer machines with MS/MS are measuring low enough levels without the need to derivatise.

I was wondering if anyone here has a newer than 15 year old LC-triple quad mass spectrometer that I might be able to try my samples on? At the moment I feel like I will never get this method done with the lack of machines I have access to and unfortunately my PhD hangs on this being completed

If anyone can help that would be great!