Advia Bench Protocol  Revision #: 4.1, Feb. 2, 2006

Revised by LLP, PM

Machine Start-Up:


Samples collected are not fasted and no ice is needed for the samples.  Pipette 20 µl of 20% EDTA (see recipe below) into 1.5 mL eppendorf tubes.  Using EDTA coated microhematocrit tubes, perform a clean retro-orbital bleed and fill tube to a final volume of 250 µL.  This procedure minimizes clot formation. Any clots will result in "Aspiration Failure", and no data will be obtained.  Check for clots with a toothpick.  If any are found discard the sample.  Run samples as indicated below.  Values reported are those obtained directly from the ADVIA; no correction for the small dilution was made.

If you haven’t used the ADVIA before, please contact the Diagnostic Lab personnel at ext. 1270 for training.

Recipe for 20% EDTA:
EDTA, Tetrasodium Salt 
Fisher Lab item number BP121-500

  1. Add 20 gm of EDTA to 100 ml mouse PBS.  Shake solution until crystals have dissolved.
  2. Check that controls have been run. These are being done by the Diagnostic Lab staff.
  3. Click STANDBY button to allow ADVIA to run.
  4. Insert NAME and ID#.
  5. Click LOG ON.
  6. Click on CUSTOMIZE - SYSTEMS SET UP - SYSTEM OPTIONS - DATA OPTIONS and go to RAW DATA DIRECTORY NAME.
  7.  Type in hlb after the \.
  8. Click on START NEW FILE - click OK then click SAVE.
  9. Click on ROUTINE OPERATIONS - MANUAL SAMPLE ID.
  10. Click on PRIMER - CBC/DIFF- 800 MOUSE. This will allow you to run the primer through ADVIA to "wet" the system.  The primer being used is called Sheath Reagent and is located on the right side of the ADVIA.  Wait for the "READY TO USE" signal before introducing the primer. After the primer has been introduced to ADVIA, change PRIMER to PATIENT and change CBC/DIFF to CBC/DIFF/RETIC.

 

Running Samples:

  1. Type in the sample ID number and click OK to send that ID up to the NEXT SAMPLE field. Always wait for the computer to tell you "READY TO USE" before introducing blood sample or you'll lose your sample.
  2. Open the eppendorf lid and slide it under the open tube wand.
  3. Push the lever behind the open tube wand to activate aspiration of blood sample.
  4. Avoid air bubbles in the line or ADVIA will throw out sample and message will say "ASPIRATION FAILED". ADVIA will chime when it has collected sufficient sample. It takes about 1 minute for the sample to run before you can introduce the next sample. Be sure to put in the proper ID. ADVIA will pop up a colored copy of printout on the screen.
  5. Click HIDE to return to MANUAL SAMPLE ID screen. Hard copy will print automatically.
  6. Begin next sample and repeat above procedure.

Data Collection:

  1. Push Ctrl and Esc buttons at the same time.
  2. On left side of screen, click on +Bin - click on +Export Data.
  3. On right side of screen, scroll all the way down to the bottom and left click on the last file.
  4. Click on highlighted file with right click and hold.
  5. Drag file to top of left side to 3.5 floppy disk and click COPY HERE.
  6. When procedure is complete, exit out of screen - click LOG OFF - click STANDBY. This prepares ADVIA for the next user.
  7. Remember to sign out on the notebook so that the Diagnostic Lab knows how many samples you ran.