01: Sample Handling & Interpretation

Component in a red top tube

None :)

Purpose of red top tubes

To clot and spin down blood to pull off the liquid that remains

What liquid is pulled from a red top tube

Serum

What tests can a red top tube be used for

• Chem panels

• Serology tests

• Anything else that needs serum

Crucial step if sending out labs in a red top tube

TRANSFER THE SERUM TO A DIFFERENT TUBE

What happens if you don’t pull the serum out of a red top tube you sent out

The RBCs will keep metabolizing and drop the glucose. You may also see hemolysis

Normal color of serum in small animals

Clear

Normal color of serum in large animals

Yellow

Component in a tiger top tube

Gel

Purpose of the gel in a tiger top tube

Separates the clot and the serum so you don’t have to transfer it

What tests can a tiger top tube be used for

• Chem profiles

• Gel may interfere with the results of some special tests

Component in a purple top tube

EDTA

EDTA mechanism

Anticoagulant that chelates Ca⁺⁺, interfering with most enzymes

What is crucial when filling a purple top tube

• No clots!!!

• Avoid hemolysis- don’t force the blood

• Don’t underfill

Problem that arises when you underfill a purple top tube

EDTA is hypertonic, and will make the RBCs shrink, artificially lowering the HCT

What tests can a purple top tube be used for

• CBC

• Blood smear

• Cytology

Why is EDTA a good anticoagulant to use for a blood smear

EDTA preserves cell morphology

Why should you rock a purple top tube before running a test

Erythrocyte sedimentation

What is a factor that influences erythrocyte sedimentation rate

Inflammation

What is the difference between serum and plasma

Plasma has clotting proteins, serum is collected from clotted blood

What tests can a purple top tube be used for

• CBC

• Chemistry profile*

What chem panel values can you NOT look at with a purple top tube and WHY

• Ca⁺⁺: EDTA is a calcium chelator

• K⁺: EDTA is a potassium salt

• Liver enzymes: may be low because they depend on calcium

Component in a greed top tube

Heparin

Heparin mechanism

Anticoag that activates antithrombin (AT), which is a natural anticoagulant in the blood

Unique side effect of heparin on blood samples

Causes platelet clumping

Handling considerations for green top tubes

Same as EDTA tubes: no clots, no hemolysis

What test can a green top tube be used for

• Chem profile (all of it!)

• CBC in birds and exotics

Why do you use a green top tube instead of a purple top tube for bird/exotic CBCs

EDTA causes hemolysis in some exotics

Component in a blue top tube

Citrate

Citrate mechanism

Reversibly chelates calcium

Important practical point when using blue top tubes

You have to fill it with the right ratio of blood

What happens if you don’t put enough blood in a blue top tube

The coag time gets artificially longer

What tests can blue top tubes be used for

• PT/PTT

• Von Willebrand facto (another hemostasis test)

How is a clot formed when running coag tests on a sample in a blue top tube

Calcium is added, overcoming the reversible anticoag

Component in a fluoride/gray top tube

Oxalate and NaF

Oxalate mechanism

Anticoag that chelates Ca⁺⁺

NaF mechanism

Inhibits glycolysis (metabolism in general)

Handling considerations for a fluoride/gray top tube

Same as EDTA: avoid clotting, hemolysis

What tests can a fluoride/gray top tube be used for

Glucose or lactate measurements that won’t be spun down and run immediately

Component in an ACT/gray top tube

Diatomaceous earth

Diatomaceous earth mechanism

Stimulates blood clotting

What tests can an ACT/gray top tube be used

Point of care hemostasis testing, useful for when you don’t have the fancy equipment for a PT/PTT

Common case where an ACT/gray top tube can offer informative diagnostics

Rodenticide poisoning

Common sites for blood collection

• Vein

• IVC

3 common issues that can be evaluated by gross analysis of serum

• Icterus

• Hemolysis

• Lipemia

Icterus cause

Bilirubin buildup, commonly from liver dz

Hemolysis cause

Dz or forcing blood through the collection needle

Lipolysis cause

• Postprandial (normal)

• Certain dz and breeds

Why are serum issues less than ideal when running a sample

Chem tests use optical density, and those serum issues may interfere with the light

Dichotomous test

Two results: yes or no, normal or abnormal

Continuous test

Numbers that exist on a scale

Sensitivity

Ability of a test to catch the disease in an affected animal

What does it mean for clin path if a test is not very specific

The value can be normal, but the patient may have the disease

• Think kidney values for a pet with CKD

What do you trust in a highly sensitive test

The negative result

Specificity

The ability of a test to categorize a nonaffected animal as normal

What does it mean in clin path if a test is specific

An abnormal result is caused by a fewer amount of things

What do you trust in a highly specific test

The positives

Precision

The ability of a test to give you the same or similar result when you repeat it

Coefficient of variation

The expected and accepted amount of variability present in repeated tests

When is the coefficient of variation a clinically relevant concept

If you are doing repeat testing (tracking progress), knowing what the normal amount of variation is lets you know if a change in a value is significant or the result of random variation

Accuracy

Is the result the test gave correct

Why is it important to calibrate your analyzers

Incorrect results an be worse than no results

T/F: every abnormal value is a problem you have to fix

False, some of them are vague and you can discard

Reference interval

The range of numbers that will include 95% of clinically normal patients

If reference intervals can be wrong, what is the best way to determine if a patient has normal or abnormal values

Compare current results to the reference range and any prior results from the pet

Key to interpreting an analyte on bloodwork

Know how the thing is produced and how it is cleared from the blood

If bilirubin has increased, what could be the problem

• Increased production (hemolysis)

• Decreased clearing (liver dz, biliary obstruction)

Key point when looking at the magnitude of an abnormality on bloodwork

Different diseases cause different degrees of change, and knowing this can help you narrow down your differentials

Liver enzymes that asses hepatocyte condition

ALT and AST

Liver enzymes that assess cholestasis (biliary obstruction)

ALP and GGT

Liver values that assess function

• Albumin

• Cholesterol

• BUN

• Bilirubin

• Bile Acids

• Ammonia