Lab Two

• Preparing samples for ProCyte DX:

  1. Use an EDTA tube that can be secured in the adapter.
  2. Use a syringe or vacuum collection system
  3. Draw the sample and transfer, if necessary
  4. Invert tube to mix
• Running samples in a ProCyte DX:

  1. Identify the patient and initiate the sample in the IDEXX VetLab station
  2. Place the sample in the analyzer
  3. Press the start button.
• Running a rinse in a ProCyte DX:

  1. Prepare five percent bleach solution up to one week before running
  2. Set aside 25 minutes to complete the procedure
  3. Follow instruction in operator's guide.
  4. The removes contaminants and checks analyzer performance.
• Maintaining a ProCyte DX analyzer:

  • Run the rinse procedure once a month
  • Run a quality control once a month
  • Change the reagent kit when low, empty or expired
  • Clean the fan filter once a month
• Running a quality control for a ProCyte DX:

  1. Use a vial of e-CHECK (XS) maintained at room temperature for at least 15 minutes
  2. Set the QC adapter in the sample drawer and insert the e-CHECK (XS) vial
  3. Follow the instruction in the operator guide
  4. This run ensures the reliability of the analyzer and reagent system
• Changing reagent in a ProCyte DX:

  1. Use the easy to pull tabs to open the new kit
  2. Fold open the top cover and do not alter the order of reagents
  3. Avoid damage to the Quick-Connect top when removing from and placing in bottles
  4. Follow the detailed instruction in the operator guide
• Cleaning the fan filter in a ProCyte DX:

  1. Turn off the analyzer and open the right side cover
  2. Remove and vacuum the fan filter
  3. Replace the fan filter and close the cover
• Preparing a whole blood sample with a catalyst lithium heparin whole blood separator:

  1. Remove the green cap from the separator to prepare it for sample collection
  2. Immediately after sample collection, in order to avoid clotting, dispense 0.7 cc of untreated whole blood into the separator using an untreated syringe with the needle removed.
  3. Use the fill line on the separator to ensure proper fill volume.
  4. Gently swirl the separator at least five time to mix the sample and the anticoagulant
  5. Do not invert or shake
  6. Ensure the cap is removed before loading into the analyzer
• Heparinized samples can be used in the lithium heparin whole blood separator except when running feline AST, LDH, or CK because that could falsely elevate results.
• Preparing a plasma chemistry sample:

  1. Use the appropriate lithium heparin tube
  2. Do not use EDTA or sodium heparin
  3. Use the appropriate sample collection device
  4. Draw the sample gently and transfer is necessary. Use the correct blood to lithium ratio
  5. When using an evacuated tube, allow the sample to draw naturally into the tube by vacuum.
  6. Gently invert the sample for 30 seconds to mix
  7. Within 30 minutes of collection, centrifuge the sample
  8. Immediately after, transfer 300 microliters of the sample to a Catalyst sample cup.
  9. Do not aspirate cells during plasma collection
• Preparing a serum chemistry sample:

  1. Use the appropriate serum tube
  2. Use the appropriate sample collection device
  3. Draw the sample gently and transfer if necessary
  4. Let the sample clot for a minimum of 20 minutes
  5. Within 45 minutes of collection, centrifuge the sample
  6. Immediately after centrifugation, transfer 300 microliters of the sample to a Catalyst sample cup
  7. Take care to not disturb the clot during serum collection.
• Preparing a urine sample for UPC ratio:

  1. Once you have obtained the urine sample through cystocentesis, a catheter, or free catch method, transfer the urine sample to a disposable sample tube
  2. Centrifuge the sample
  3. Use a transfer pipette to transfer 300 microliters of supernatant urine to a Catalyst sample cup.
  4. Dispense 300 microliters of Catalyst urine P:C diluent to a Catalyst sample cup.
• When running a lab, you need to be familiar with:

  • The types of analytical instruments
  • The variety of testing procedures used
  • Rationale underlying analyses
• Most chemical analyses require whole blood, plasma, or serum.
• Plasma:

  • Fluid component of blood
  • Made up of 90 percent water and 10 percent dissolved constituents
• Getting a plasma sample:

  1. Draw the blood sample
  2. Place in the appropriate anticoagulant tube, like lithium heparin
  3. Rock blood gently for 12 minutes
  4. Place in the centrifuge
  5. Use a pipette to remove the plasma layer
  6. Transfer to a container
  7. Proceed with testing, or refrigerate/freeze
• Serum


  Fluid from blood that has fibrinogen, a plasma protein, removed.
• Getting a serum sample:

  1. Collect a blood sample
  2. Place in a container with no anticoagulant
  3. Red top or tiger top tube
  4. Allow blood to clot for 20 minutes
  5. Do not refrigerate blood until it is clotted
  6. Gently separate clot from the tube by running a wooden applicator stick around the wall of the tube
  7. Centrifuge for 10 minutes
  8. Remove serum from the clot using a pipette
  9. Proceed with testing, or refrigerate/freeze
• Factors influencing blood chemistry:

  • Hemolysis
  • Alters the makeup of the serum or plasma sample
  • Chemical contamination
  • Improper labeling
  • Improper sample handling
  • All measurements should be handled within an hour of collection
  • Samples should not become too warm
  • Patient influences
• Chemical contamination of blood chemistry:

  • Not necessary to use sterile tubes, but they should be chemically pure
  • Completely rinse detergents from tubes before reuse.
• Tubes should be labeled with:

  • Date
  • Time of collection
  • Patient name
  • Doctor name
• Samples should be taken from a fasted patient otherwise glucose levels and turbidity of the sample can be affected.
• Endpoint analysis:

  • A product is formed from enzymatic action that interacts with a reagent to produce a color complex
  • The amount of light absorbed by this color complex is proportional to the amount of color complex present, which indirectly reflects the concentration of enzyme in the sample
  • The more intense the color, the more product that was produced because there was more enzyme present.
• Factors that affect enzyme activity:

  • Inhibited by:
  • Low temperature
  • Dehydration
  • Ultraviolet light
  • Salts of heavy metals, like copper and mercury
  • Accelerated by:
  • High temperature
• Tips for samples:

  • Store sample in a cabinet above the blood machine and leave them there all day, and only remove when the clinic is closing.
  • Prevents you from having to draw extra blood
  • Keep samples dark and at a cooler temperature
• When preparing a blood film, use fresh blood that is less than 24 hours old because specimen deterioration occurs with prolonged storage
• Blood film technique:

  1. Place a small drop of fresh, well mixed anticoagulated blood on a clean glass slide about two centimeters from one end of the slide
  2. Place a clean glass spreader slide in front of the drop of blood at about a 30 degree angle to the film slide
  3. Increase the angle for low Hct/anemia blood
  4. Decrease the angle for high Hct/dehydration/polycythemia blood
  5. Back the spreader slide into the drop of blood
  6. Let the blood spread along the contact line between the two slides until it covers 3/4 of the width of the slide
• Blood film technique part 2:

  5. With steady and seamless movement, move the spreader slide down the entire blood film slide, maintaining the angle without lifting the spreader slide. Blood will follow the spreader slide, placing a thin film on the other side

  A, The blood film should be three to four centimeters in length and the shape of a thumb print

  6. Let the slide air dry or use a fan to avoid air drying artifacts

  7. Stain the slides and allow to air dry or use a fan
• What to look for in a blood film:

  • Visual
  • A thumbprint appearance
  • Presence of a feathered edge
  • Microscopic screening ( 4X, 10X, or 20X objective)
  • Presence of a monolayer
  • Minimal to no stain precipitate
• What to avoid on a blood film:

  • Visual
  • Uneven staining
  • Incomplete and asymmetrical feathered edge
  • Microscopic evaluation
  • Specimen over 48 hours old
  • Uneven film
  • Lysed cells
• In blood films, do not:

  • Use a dirty or chipped spreader slide
  • Go too fast
  • Hesitate with the spreader slide
  • Use a drop of blood that is too small
  • Move so quickly that the blood doesn't have a chance to move across the spreader slide
  • Use slides contaminated with dirt or grease, as elevated blood lipids have a similar appearance
  • Use uneven pressure on the spreader slide
  • Allow the drop of blood to begin to dry