Navigating Spurious Results on CBC/Chemistry

“Treat the patient, not the numbers”, was the first lesson I learned during my clinical pathology course in veterinary school.

Most of the time, lab results from automated analyzers reflect the clinical picture of our patient. At times though, we are faced with results that leave us scratching our heads, or worse, derail us in our diagnostic path.

As an industry veterinarian specialized in diagnostics, I have seen a lot of technological advances in the clinical lab field. While these advances are usually helping us gain time and more information on our patients, it comes also with a new set of issues - namely discrepant or hard to interpret results.

Interpreting results from any analyzer requires a good understanding of the technology behind it. This is what I want to share with you today: the common discrepancies that every veterinarian should be aware of when starting their career in clinical practice.

Laboratory sample processing errors can be divided into three categories: preanalytical, analytical, and postanalytical errors. Automation has significantly reduced the errors occurring in the analytical and postanalytical phases, whereas the preanalytical stage still has a long way to go as it is mostly dependent on manual labor.

Examples of errors that arise in the preanalytical phase include:

• Incomplete sample labeling.
• Improper collection technique (clotted, hemolyzed or insufficient sample).
• Sample contamination when collecting from catheter (diluted or contaminated).
• Wrong tube / anticoagulant.
• Delayed mixing of blood with anticoagulant.
• Improper mixing of blood before presenting sample to analyzer.
• Delayed removal of serum or plasma clot.
• Delayed sample analysis.
• Improper sample storage.
• Inadequate temperature of sample before analysis.
• Inadequate patient preparation (not fasted, medication).
• Interference substances in sample.

In the clinic, pre-analytical errors can be minimized by implementing a rigorous protocol for technicians and doctors to follow:

• Best methods to draw blood on our patients.
• Information given to pet owners to prepare patients for lab work (i.e., fasted, medication given/withhold, ...).
• Workflow of laboratory test ordering.
• Collection and storage (i.e., samples and tubes patient identification, proper tube collection and adequate storage if required).

The following analytical errors can happen during the actual performance of the test:

• Operator errors (improper reagent, improper sample handling, or incorrect analyzer use).
• Reagent issues (out-of-date reagents, improperly stored reagents).
• Analyzer malfunction (improper maintenance and/or calibration, aging of instruments).

CBC and Chemistry at the point-of-care

CBC and chemistry analyzers at the point-of-care vary in technology. Some analyzers require more maintenance and regular calibration than others. If the clinic cannot allocate the time and resources to properly maintain and calibrate their in-house equipment, reviewing protocols for in-house vs send out lab samples analysis may be warranted.

The Complete Blood Count CBC

When talking about CBC specifically, EDTA tubes have a set fill volume that ensures the best anticoagulant concentration, and both under- and overfilling can result in incorrect complete blood cell count readings.

Underfilling the tube, i.e., having a greater ratio of anticoagulant to blood, may shrink red blood cells artificially. This causes a lower MCV (mean cell volume), and in turn lowers HCT (hematocrit) and increases MCHC (mean corpuscular hemoglobin concentration) because these are calculated values on most analyzers.

Overfilling an EDTA tube will risk the sample to clot and invalidate results on CBC automated analyzers. While micro clotted samples can be run on automated analyzers, they may falsely decrease the number of platelets because they are involved in micro clots. Inadequate mixing of the tube after collection or prior to testing may cause similar abnormalities.

Platelet clumping

Platelet clumping is a common reason for a reduced number of platelets on a CBC. Cats’ platelets are more reactive than dogs and we witness this phenomenon more frequently in that species. Why does it occur? Frequently an artifact of platelet activation during collection or handling at time of venipuncture, platelets clumps can be identified by making a blood smear and looking for large clumps in the feathered edge or smaller clumps in the monolayer. This extra step is important prior to any surgical procedure or attempting any diagnosis.

Spurious pancytopenia

On in-house analyzers, results showing reduced numbers in all cell lines, called pancytopenia, should always prompt the clinician to make a blood smear to rule out an insufficient volume of blood picked up by the analyzer.

If your clinic staff doesn’t have the time to go through this extra step, you may consider sending out your CBCs to the reference lab, where experienced lab technicians will proceed with a manual evaluation of a blood smear.

Sample Interferences and the CBC

Lipemia

Interferes with hematology results through the light scattering mechanism. This will artificially increase hemoglobin and hemoglobin related indices (MCH and MCHC). If lipemia is severe, it can sometimes interfere with platelet count, as the large lipid droplets get falsely counted as platelets.

Hemolysis

Interferes with hematology results through the breakage of the RBCs and light scattering. The count of RBCs will be lower, which in turn will affect the Hematocrit HCT result, which is a calculated value on most CBC analyzers.

Icterus

No interference has been reported from icterus in hematology.

Chemistry Panels

A common chemistry abnormality is EDTA contamination of blood samples. It is easily recognized by marked hyperkalemia and hypocalcemia. The anticoagulant potassium EDTA is used for CBC analysis, and it is not uncommon to see mild contamination of the chemistry tube from the needle used to transfer blood in the tubes. There is no fix for this and a repeat analysis with a new blood sample will be required.

Sample Interferences and the Chemistry

Hemolysis

When the serum or plasma display a reddish color, hemolysis is most likely present. Hemolysis releases free hemoglobin that may interfere with the color reading of spectrophotometric assays. By breaking down red blood cells, the contents of the cells will artificially increase certain values such as potassium, LDH (lactate dehydrogenase), AST (Aspartate dehydrogenase) and CK (creatine kinase). Knowing that hemolysis has occurred and knowing roughly the severity of it will help the clinician interpret the results of these analytes accordingly.

Lipemia

Lipemia causes serum and/ or plasma to appear milky white when the triglyceride concentration exceeds 300 mg/dl. Lipemia interferes with chemistry tests by the following mechanisms:

• Light scattering: some results may be falsely increased due to increased absorbance readings, for example total Bilirubin.
• Volume displacement: Fat in the serum displaces an equal volume of water. Because some analytes (sodium and chloride) are dissolved in water, not in lipids, this may falsely decrease levels of sodium or other substances, while physiologically the patient has no deficiencies.
• Lipemia causes red blood cells to be more fragile and may cause additional hemolysis to the sample.

Icterus

Icterus is the yellow appearance of serum or plasma when bilirubin levels are elevated. The color change affects the chemical reaction to detect creatinine and total proteins concentrations, which both will be decreased respectively when high bilirubin concentrations are present.

In conclusion, proper sample collection and handling techniques are essential to avoid preanalytical laboratory errors and obtain reliable results both at the point-of-care and at the reference laboratory.

In-clinic testing has the advantage of rapid turnaround time and minimizes preanalytical errors associated with transportation and delayed analysis of samples. Some in-clinic testing remains necessary for emergency cases. The disadvantages include the time and expertise required of clinic staff to assure quality and reliable results through maintenance of reagents and equipment, quality assurance through calibration and quality controls, troubleshooting analyzer issues, preparation and microscopic analysis of blood smears.

Sending samples to the reference laboratory may be a viable option for clinics lacking resources to assure quality results at the point-of-care. Ellie Diagnostics offers more cost-effective testing, experienced personnel specialized in laboratory testing, better quality assurance, better detection of laboratory errors, access to specialists to review results and a wider range of tests.

Ariane Kaplan DVM, MPH

Dr. Kaplan earned her DVM and master’s in public health from Tufts University in Massachusetts, followed by a small animal rotating internship at VCA in Phoenix, Arizona.

She practiced small animal medicine and emergency medicine before becoming a Diagnostics Professional Services Veterinarian 6 years ago. She is currently the Director of Veterinary Relations at Ellie Diagnostics.

In her free time, she consults with her 4 misfit cats on good manners and how to live together in harmony.