Can’t Communicate with my Instrument (GC or LC)!

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There’s nothing like  preparing your samples and starting chemstation, only to see “instrument offline”.  There are some basics that can be checked that may be the solution if you are having this problem.

IP addresses:

You always want to start here.  A 68/7890 GC, as well as 1100-1290 LCs use LAN communication for the most part.  The main idea being, is the ip address of the instrument and the ip address of the pc compatible?  You want the ip address to match up until the last set of numbers.  For example:  if the gc is 10.1.1.101, the pc needs to be 10.1.1.x.  If the LC is 192.168.254.11, the pc needs to be 192.168.254.x.  With “x” being any digit from 1 to 255.

How can I check the ip address of the GC?  This is pretty simple, on the majority of these GCs you can find out by hitting options, then communication on the gc front panel.  From here you can see what the address currently is, and you can also change it as well.

The LC is not as straight forward, but the default is 192.168.254.11.  You can also connect the gameboy to the module and view/edit the ip address.  If it’s not the default, and you are not sure what the address is, then you will have to go through a process of changing switches on the LAN card and setting it to default.  From there you can keep the default or change to a new address using Telnet.  I may write an article on that procedure in the future.

How can I check the ip address on the pc?  You can do this through the control panel of windows, then go to network and internet, then network connections.  You can see all your network connections on this screen.   Most people will have two connections, a “house” or company network, and a local area connection.  The local are connection is usually the one that is connected to the instrument.  Right-click on that connection, then double click on IPv4.  You will see a screen like this:

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From here you can view and change the IP address.  Make sure to keep the subnet mask: 255.255.255.0.  Hit OK to save the changes.

After determining that the addresses are compatible, the next thing you want to do is ping the instrument.  To do this you need to launch the command prompt.  In Windows 7, click on the Start key for windows, in the search bar type CMD and hit enter.  In Windows XP, hit start, and select run, type CMD and hit enter.

Once in the command prompt type: ping 10.1.1.101 (or whatever the ip address of the instrument is).  You should get 4 replies back, 100% packets sent and received.  If not, then check that the cable from the pc to the insturment is connected and good, maybe try swapping it out.  If there is a switch or hub in between make sure that port is operational.  If there are 2 LAN cards on the pc, its possible that you are connecting to the wrong one, or changed the ip address on the wrong one.  Does the LAN card on both the pc and instrument show flashing LED lights on the port?  If not it may be a hardware problem with one of the cards.

 

Configuration: 

If pinging the instrument was successful, the last piece of the puzzle is ensuring the configuration in chemstation has the proper ip address for the instrument.  If you are using a classic version of chemstation (pre-Openlab), go to start, all programs, agilent chemstation, and launch the configuration editor.  From here you can view/edit the ip address of what chemstation thinks the instrument is configured to be.  In Openlab chemstation, launch the Openlab control panel, select your instrument on the left hand side, and click on configure instrument on the top of the screen.  From here you can view/edit the ip address of what Openlab is expecting your instrument to be configured to be.

 

 

I hope these basic steps of checking/pinging the ip address have been helpful.  Please leave a comment if you have any feedback or questions, thanks!

 

 

 

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Extracting a Signal from Spectral Data

If you collected spectral data in your run and would like to extract a wavelength from that data, you can follow these steps below.

First, load your data file.  Next, select Spectra then Isoabsorbance plot.

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On the upper left box on the screen you want to change Cursor to Signal.  Then select the wavelength you would like to extract.  In this example I chose wavelength 200.  You can also specify a reference signal and bandwidth as well if needed.

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After selecting the desired signal, push copy at the bottom then hit ok.  You may get a pop-up message about temporary data, you can hit ok to that as well.  You will now see your chromatogram for the signal you extracted.

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How to Build a Calibration Table in Chemstation (with screenshots)

This will be a quick tutorial on how to build a calibration table in Chemstation, with screenshots.  The first thing that you want to do is run your standards.  Most of the time you want to have 3 levels; low, medium, and high.  You want your compound of interest to have a concentration close to the mid-range level.  For example, if my samples will typically have a concentration of 50 ng/ul, then my standards could be 25, 50, and 75 ng/ul.  Some people might have five or more levels, others might just have one level.

Next step is integration, you want to be sure that your peaks of interest are all integrated, and you can filter for unwanted peaks (possible blog post about that process).

After running your standards, and integrating, the next step would be to select calibration, and new calibration table.

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I usually leave the settings with the default and hit ok.  Now, every single peak that was integrated earlier will show up on your calibration table.  Unwanted peaks can be removed by keeping zero as the amount.

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You can tell the integrated peaks by the pink line beneath the peak and the retention time label above it.

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At this point, when you click on each line, you can see which peak you are on and give it a name and an amount.  Again, the units for amount are totally customize able.  I had a post about this earlier.

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After adding the amount, hit ok, and chemstation will delete line with zero amounts.

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The peaks now have the name that you specified above them.  Load your level two calibration run, go to calibration and select Add Level.

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Do the same thing this time, you’ll only have to add the amount for each peak with each subsequent level.  After adding all of your levels, you can look to the right of the calibration table and see the calibration curve, equation, and r value of each compound.

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The calibration table is now complete.  Be sure to select the proper quantitative settings by going to report, then specify report.  For this type of calibration table you want to select ESTD based on area.

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High Pressure in my HPLC!

Ok so no one wants to see the pressure on their HPLC higher than what it should be.  In my experience the number one cause of high pressure is a clog in the autosampler.  Followed by a clog in the purge valve frit, as the second highest cause of HPLC over pressure.  Luckily, there’s an easy way to diagnose whether or not there’s a clog in the autosampler.  This technique works on Agilent model HPLCs and Chemstation, not sure about other brands.

Turn the pump on, once you see the high pressure, right-click on the diagram of the autosampler.  Select the switch valve to bypass option (see below).  See if pressure drops immediately, if so change the needle and needle seat (or backflush).

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Note: Older versions of chemstation will only have that option available in the diagnosis tab.  I may make a tutorial on the process in the future.

What if the pressure is still high?  Try removing the column from the flow path.  If that doesn’t help, try removing the autosampler from the flow path.  If that doesn’t work, try opening the purge valve.  If opening the purge valve does not decrease the pressure, try replacing the purge valve frit.

Pro Tip:  When installing a brand new column, make note of the pressure of the system.  This is what your pressure baseline is, and will be a good reference in the future when you are not sure if your pressure is where it should be.

Chemstation Calibration Units

Pro Tip: Chemstation does not look at the units when calculating amounts.

Example:

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As you can see the amounts is in ng/ul.  If I go to calibration–> calibration settings, I can choose any unit (or non unit for that matter) I like.

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In this example I chose Aliens as my unit of choice.

amt3

As you can see, Chemstation willfully changes the units to that value.

Why is this important?  Many people think that Chemstation will convert units for you.  You can add a dilution or multiplication factor, but that will be used in the calculation of the amount regardless  of what units you choose.  Best practice would be to build your calibration table based on the units which you would like your results to be, so that by the time you print the report, you won’t need to convert any units.

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