1. Proper Sample Storage & Submission

study of disease using laboratory assays

clinical pathology

the study of blood and how the body is responding to disease

hematology

the measurement of physiologic analytes and biochemical reactions

clinical chemistry

the study of cells, exfoliative or in suspension

cytology

variably incorporates automated cell counting, protein measurement, biochemical assays, and cytology

fluid analysis of non-blood body fluids

True or false: Veterinary practitioners utilize a foundation of clinical pathology knowledge every day. The more you know, the better you’ll understand and respond to the laboratory data you collect. Veterinary clinical pathologists are specialized veterinarians employed in industry, academia, and research to guide the translation of laboratory data into a meaningful assessment of animal disease.

true

True or false: Make sure you read the labels, do not just go by color.

true

What needs to be done before you draw the sample?

know the submission requirements

What should be done when running atypical tests, changing analyzers, or changing reference labs?

pay special attention

What should be done if in doubt?

consult the analyzer manual or manufacturer or contact the lab you’re submitting to

represents an unspun, non-clotted sample, ± anticoagulant that is commonly used with point-of-care handheld analyzers in urgent care, anesthetic monitoring, or field settings and for CBCs

whole blood

fluid fraction of whole blood obtained from an anti-coagulated sample and is commonly used for bench top biochemical

plasma

fluid fraction of whole blood obtained from a clotted sample (loss of clotting factors) and is commonly used for bench top biochemical testing and is sometimes required

serum

What is used for a CBC?

whole blood

What is used for a chemistry?

plasma and serum

What is a buffy coat made up from? Will it be seen with a plasma sample or a serum sample?

WBCs; plasma

What does serum equal?

plasma - fibrinogen

What can lead to pre-analytical errors?

poor blood collection technique, storage, and submission

What can be the consequences of traumatic venipuncture?

may introduce hemolysis and or artificially reduce platelet counts

How should blood samples be transferred into tubes? Why?

promptly; blood starts the clotting process as soon as it starts being drawn

Should purple tops be filled first or last? Why?

last if possible; avoid EDTA contamination of samples intended for biochemistry

When filling the collection tubes, what should be avoided? What should be done instead? Why?

plunging the syringe; allow the tube to fill via vacuum pressure; excess pressure introduces hemolysis

When it comes to anticoagulant tubes, what needs to be ensured?

proper volume and prompt, gentle tube inversion for mixing

What should be done with plasma or serum obtained for biochemical testing if it is not going to be immediately used?

it should be properly stored

At minimum, what is the proper storage for plasma or serum?

refrigerated or immediately processed for shipment on ice

What can excess heat with plasma or serum cause?

erroneous values

When shipping your blood sample, what do you need to make sure?

the sample is secure, cushion any extra space, and waterproof paperwork

What else is an option for plasma and serum samples? When is this commonly used?

freezing; commonly used for long-term storage in clinical research or for certain auxillary tests such as send-out coagulation testing

What should not be done with CBC samples?

centrifuged or frozen

What is best to be done with CBC samples?

best to run CBC samples ASAP

If the CBC is refrigerated, how is it generally acceptable? How long will many labs store them up to?

if refrigerated then brought back to room temperature, generally acceptable within 72 hours; 7 days

What tubes are anticoagulant tubes?

• EDTA (purple top)

• heparin (green top)

• citrate (blue top)

Which tubes are plain tubes?

• red top

• white top

tube with K+ and Na+ EDTA variants that prevent coagulation by chelating calcium and is gentler on cells

EDTA, purple top tube

For what blood sample is EDTA, purple top tube used for?

CBC and whole blood

inhibits coagulation by potentiating antithrombin

heparin, green top tube

For what blood sample is the heparin, green top tube used for?

plasma biochemistry

reversibly prevents coagulation by weakly chelating calcium

citrate, blue top tube

For what blood sample is citrate, blue top tube used for?

coagulation testing

species dependent clotting time prior to centrifugation

red-top

What is a red-top tube used for?

serum biochemistry

miscellaneous use for sample storage

white top

What are serum separator tubes also known as?

tiger top tubes

What do serum separator tubes contain?

a gel with intermediate density between cells and serum or plasma

True or false: Upon centrifugation of a tiger top, the fluid fraction will be separated from the cell fraction, preventing leeching of certain analytes into the cell fraction. If not using a tiger top, immediate separation of fluid from cells is necessary.

true

Some tests, such as phenobarbital measurement, are falsely ________ from fluid fraction contact with the gel separator, so check sample requirements.

decreased

With the gold top tube, how long should it be allowed to clot for? When should it be centrifuged before?

30 minutes; 2 hours after clotting

results in free hemoglobin and subsequent red discoloration that may be in vitro or in vivo intravascular

hemolysis

results in lactescent appearance and is commonly observed in post-prandial samples and why small animal patients should ideally be fasted for a blood draw

lipemia

represents elevated bilirubin and yellow discoloration that use species specific rubrics, as large animal plasma and serum naturally have a moderately yellow appearance

icterus

What can significant changes to plasma and serum appearance cause?

interference with CBC and or clinical chemistry measurements

In vitro hemolysis falsely decreases what values?

• packed cell volume (PCV)

• hematocrit (HCT)

• red blood cell count (RBC)

What is the affect of in vitro hemolysis on mean corpuscular hemoglobin concentration (MCHC)? What will be unaffected?

falsely increased; MCV

________ ________ from intravascular hemolysis may be falsely counted as ________, ________ PLT.

ghost erythrocytes; platelets; increasing

What becomes difficult to read due to in vitro hemolysis?

refractometric protein

What is the affect of severe hemolysis on electrophoresis?

can cause beta globulin spikes

What is the affect of hemolysis on chemistry?

S
I
M

• spectrophotometric interference and inhibition of chemical reactions

• increase in intra-RBC analytes

• minimal to mild increase in creatine kinase (CK) enzymatic activity

In hemolysis, which intra-RBC analytes are specifically increased?

P
P
A
L
M

• potassium

• phosphorus

• ALT

• LDH

• Mg

What is the effect of lipemia on CBC?

F
L
F

• falsely increases HGB and subsequently MCHC

• large lipid aggregates may be falsely counted as platelets, increasing PLT, or potentially leukocytes, increasing WBC

• falsely increase refractometric protein

What are the affects of lipemia on chemistry?

S
P
S

• spectrophotometric interference

• proportional decrease in Na and Cl

• slight decrease in K

What does lipemia also promote?

in vitro hemolysis

What can be done with samples that have lipemia? Why?

can be refrigerated; precipitates out the lipid, allowing collection of less lipemic serum/plasma

What is the affect of icterus on CBC?

little to no effect

What is the effect of icterus on chemistry?

falsely decreases biuret total protein and creatinine