Chromatography Forum

i was wondering what is the proper way of using a syringe when injecting through a septa into a inlet port. We have two syringes, one of them is a plunger type.

Lets say I use the syringe to take a sample from a vial, my goal is to take 2 ul. When I plunge the syringe into the vial, do I just go up to 2 ul or do I go up and down few times and then slowly go to 3 ul and take the syringe out and push the remaining liq. out to the desired level.

I use this technique for manual injection

do I go up and down few times and then slowly go to 3 ul and take the syringe out and push the remaining liq. out to the desired level.

except that I first suck some sample into the syringe then eject this first aliquot into a waste container.

http://www.hamiltoncompany.com/download ... %20Use.pdf

This "Care and Use Guide" from Hamilton Comapny may be of worth to you.

I just go up to 2 ul or do I go up and down few times and then slowly go to 3 ul

I follow the above but I dispense the extra volume with the syringe still in contact with the sample/liquid. If you remove the syringe and expel the extra, how do you remove the extra?

There are several schools of thought (we are analytical chermists after all) regarding which technique gives the most reproducible results and I'll mention a few below.

1) Pull the needle out of the solution and dispense the extra volume with the needle in contact with wall of sample vessel. The only question here is, do you lose additional sample due to capillary action/surface tension.

2) With the needle still in the solution, dispense the extra volume then remove the needle from the solution. The question here is, does additional liquid remain on the needle.

Additonally, after getting the desired volume in the syringe some propose to pull up a few uL of air (so as to not remove sample thru adsorption onto the wipe) and wipe off the needle.

3) A further variation (probably way more than you wanted to know at this point) is to flush the syringe with clean solvent then pull a microliter of solvent into the syringe. Then pull a microliter or 2 of air followed by the desired volume of sample. You can pull the sample up a few times to rinse the syringe, just dont expell all liquid, i.e. leave some of the air and all of the clean solvent in the syringe. This variation, some argue, ensures better precision by rinsing the sample out of the syringe during the injection process and reducing variability due to evaporation from the needle while in the injection port (time dependant). Just be sure the total volume of solvent (clean + sample) doesn't exceed the vapor volume of your inlet liner.

Either way, as long as you follow one of the techniques consistently your results should be consistent.

Also remember that whatever method you use to make the injection ( for both standards and samples), if you introduce an "error" and the "error" is always consistent then your data will be be constitent and accurate.....the only other thing I usually do is to take the first injections and eject them into waste, it will not contaminate your dample from the others and it will not introduce waste into your vial.

I just go up to 2 ul or do I go up and down few times and then slowly go to 3 ul

I follow the above but I dispense the extra volume with the syringe still in contact with the sample/liquid. If you remove the syringe and expel the extra, how do you remove the extra?

There are several schools of thought (we are analytical chermists after all) regarding which technique gives the most reproducible results and I'll mention a few below.

1) Pull the needle out of the solution and dispense the extra volume with the needle in contact with wall of sample vessel. The only question here is, do you lose additional sample due to capillary action/surface tension.

2) With the needle still in the solution, dispense the extra volume then remove the needle from the solution. The question here is, does additional liquid remain on the needle.

Additonally, after getting the desired volume in the syringe some propose to pull up a few uL of air (so as to not remove sample thru adsorption onto the wipe) and wipe off the needle.

3) A further variation (probably way more than you wanted to know at this point) is to flush the syringe with clean solvent then pull a microliter of solvent into the syringe. Then pull a microliter or 2 of air followed by the desired volume of sample. You can pull the sample up a few times to rinse the syringe, just dont expell all liquid, i.e. leave some of the air and all of the clean solvent in the syringe. This variation, some argue, ensures better precision by rinsing the sample out of the syringe during the injection process and reducing variability due to evaporation from the needle while in the injection port (time dependant). Just be sure the total volume of solvent (clean + sample) doesn't exceed the vapor volume of your inlet liner.

Either way, as long as you follow one of the techniques consistently your results should be consistent.

Oops,

#3 only applies to GC

You should post this question on the GC forum. I'm sure you'll get a much bigger response.

I admittedly didn't read every word here but this is something that people that use my syringes (damn them!) always forget:

Pull up slowly, push out quickly. If you've ever seen an autosampler work, it pulls up agonizingly slowly and consistently, but then when it pushes down, it's a quick, jerky movement.

I flush the syringe with a bunch of slow pumps, then I pull it above the level of sample (but still in the vial) and pull up about 1 uL of air. Then I put it in the sample and pull up what should be 1 uL. I move it above the sample and then pull in another 1 uL of air so the sample is sandwiched by air. I can then examine it closely to see if I have as close to 1 uL as I can get. If its not right I just start over with the pumps.

If you get good at this you can reduce error pretty well.

Also, if you're making a manual injection when I pull the needle out of the septa I keep my arms locked in position and "lift" it out by stepping up on my tip toes. Otherwise without noticing you will pull back as you pull up--tearing the septa.

99% of our injections are with GC autosampler. Get that if you want precision and accuracy.