Clinical Biochemistry Flashcards
define clinical biochemistry
clinical analysis of body fluids for the diagnosis, therapy and prevention of diseases
what can be sampled for biochemistry?
usually serum/plasma but can be urine and other body fluids (e.g. pleural fluid)
what body systems/parameters does a typical biochemistry panel include?
liver kidney proteins electrolytes glucose lipids
what are panels separated into?
organ or system
what liver parameters are typically assessed on a biochemistry panel?
ALT, AST, GLDH, ALP, GGT, Bilirubin, bile acids
what liver parameters demonstrate liver damage?
hepatocellular damage/ leakage enzymes - ALT, AST, GLDH
what level of liver damage does ALT show?
mild
what level of liver damage does AST show?
necrosis
what liver parameters demonstrate bile accumulation?
cholestatic enzymes - ALP, GGT
what substances can be measured on a biochemistry panel that are conjugated and excreted by the liver?
bilirubin
bile acids
what are the main components assessed when looking at the liver?
enzymes and function markers
levels of what substances produced in the liver are tested on a biochemistry panel?
cholesterol, urea, albumin, coagulation factors
what are liver parameters measured in?
level of increase above the reference interval (e.g. 10x)
what enzyme level can be elevated in dogs without cholestasis?
ALP is increased by steroids and phenobarbital
what can AST levels be increased by as well as liver damage?
muscle damage
what is measured to assess kidney function?
urea
creatinine
what is measured to assess protein levels?
TP, albumin
what is measured to assess electrolyte levels?
Na, K, Cl, Ca, PO4
what is measured to check blood sugar levels?
glucose
what is measured to assess lipid levels?
triglycerides
cholesterol
what does quality control ensure?
results are reliable
what should be involved in ensuring quality control?
set up of machines maintenance and cleaning of machines interpretation of results checks to ensure values are acceptable (control test) recording
what does the level of cleaning of biochemistry machines depend on?
level of use - if used frequently then may not need to be cleaned every time
what should be checked during the maintenance of biochemistry machines?
temperature and pH of reagents and that they are up to date. This ensures reactions happen properly and enzymes have optimum working conditions
why is quality control of biochemistry results so important?
decisions may be made based on wrong values - even euthanasia
results may have been wrong for a prolonged period of time without anyone being aware
what are the 2 main variables which affect test results?
biological
analytical
what are the 2 key biological factors which can affect test results?
inter-individual
intra-individual
what are the 3 analytical factors which can affect test results?
pre-analytical (before)
analytical (during)
post-analytical (after)
what variables will affect the result most pre-test?
biological and pre-analytical
what are inter-individual variables?
inherent differences between groups of animals due to the effects of species, breed, age and sex
what is an example of a common inter-individual variable associated with species?
cats have lower PCV than dogs
what are common inter-individual variables associated with age?
growing dogs have higher Ca, phosphate, and ALP but lower total protein concentration than adult dogs (this will also differ between breeds)
what are intra-individual variables?
transient differences in the same animal due to environment/external factors
what are examples of environmental/external factors that will form intra-individual variables?
diet, excitement, reproductive status, drugs, method of sampling
what factors should be minimised?
intra-individual
how can intra-individual diet factors be minimised?
all patients fasted for 8-12 hours pre biochemistry
what are the main pre-analytical factors that affect biochemistry results?
poor sampling/type of sampling haemolysed, lipaemic or icteric plasma wrong anticoagulant wrong anti-coagulant/blood ratio wrong tube transport of sample storage contamination with EDTA
in what order must anticoagulant/blood ratios be respected?
clotting
haematology
biochemistry
how should samples be stored/transported?
refrigerated not frozen unless serum/plasma are separated
how can contamination with EDTA be avoided?
don’t touch tube when transferring sample from the clotting test tube as it will ruin biochemistry tests
what method may give a falsely low reading during enzyme testing?
kinetic (enzymatic) assays instead of end point assays
what is the difference between end point and kinetic (enzymatic) assays?
kinetic (enzymatic) assays measure activity rather than amount of enzyme at the pre-determined end point
why may kinetic (enzymatic) assays give a false low?
if there is high amount of enzyme which leads to substrate depletion and so reduced enzyme activity
how can false lows in kinetic (enzymatic) assays be retested?
diluting solution
what are reference intervals?
most prevalent value in the population
what percentage of the population fall inside reference intervals?
95% of population
1 in 20 will have readings outside RI and be healthy
as as well between patients where will reference intervals vary?
between biochemistry machines
describe the pattern of changes in liver enzymes due to acute hepatic injury
ALT is most sensitive and shows initial increase, will also be the last to decrease
AST is less sensitive as it is found within the mitochondria of hepatocytes
describe the pattern of changes in liver enzymes due to cholestasis
cholestatic enzymes will increase to deal with increase bile presence (blockage preventing drainage)
if issue is prolonged then bile will begin to cause damage leading to increase in damage enzymes
what is an increase in bilirubin known as?
hyperbilirubinaemia
what does hyperbilirubinaemia lead to?
jaundice
what is bilirubin?
breakdown product of heamoglobin in RBC
where is bilirubin processed and excreted?
processed in liver and excreted in the faeces through bile
what are the 3 causes of hyperbilirubinaemia?
pre-hepatic: increased RBC breakdown
hepatic: decreased bilirubin processing (e.g. liver disease)
post-hepatic: decreased bilirubin excretion (e.g. bile duct obstruction)
where are bile acids produced and excreted?
in the liver and excreted into small intestine via bile duct
where are bile acids reabsorbed?
in distal small intestine
what is reabsorption of bile acids known as?
enterohepatic recycling
what are the 3 main reasons for bile acid elevation?
decreased hepatic function
decreased bile flow (cholestasis)
portosystemic shunt
what is a portosystemic shunt?
flow of blood through portal vein to liver enters the systemic circulation instead
what does a jaundiced sample suggest about bile acids?
they will be high
describe the bile acid stimulation test
fast overnight, take fasted sample into a plain tube, feed, take a sample into a plain tube 2 hours after feeding
what must be checked before sample collection?
tube type, volume of blood/plasma required for each test
why does the volume of blood not equal the volume of plasma?
PCV makes up around 45% of blood volume in dogs which is not usable for biochemistry
how much blood should you aim to collect when sampling?
3x the amount of plasma volume required - allows for PCV and less complaint patient
how can haemolysis and platelet clumping be avoided in sampling?
clean stick
avoid vacuum and collapse of veins by not drawing back on syringe too quickly
what tubes can be used for biochemistry tests?
plain tube/serum
heparin tube
Oxf tube
what should Oxf tubes be used for?
measurement of glucose when exact values needed as Oxf stops RBC continuing to use glucose
what is happening to glucose in plain and heparin tubes?
cells are still living and so glucose is still being consumed
what does lipaemia lead to visibly?
turbid/milky serum/plasma
what is lipaemia due to?
presence of lipids
what effect will lipaemia have on values?
will increase and decrease some values in plasma/serum due to extra lipid fractions and turbidity caused by lipids
what will ideal biochemistry systems do with regards to lipaemia?
warn of presence and grade it
does lipaemia affect hematology?
yes
what does lipaemia affect that influences results?
light transmission which will affect spectrophotometric assays)
can result in apparent dilution of normal substances in aqueous component leading to a false low (e.g. electrolytes)
what are the physiological reasons for lipaemia?
post-prandial (need fasted samples!!)
what are the pathological reasons for lipaemia?
endocrinopathies e.g. diabetes, hypothyroidism, hyperadrenocortisolism
will pathological lipaemia dissappear when fasted?
no
what does haemolysis cause?
red coloured serum/plasma due to free haemoglobin/myoglobin
what may cause haemolysis?
in vitro - pathological
in vivo - improper handling/sampling most common
what does haemolysis increase in biochemistry?
plasma/serum values of some compounds/enzymes due to increased concentration of RBC
what does haemolysis decrease in biochemistry?
plasma/serum values of compounds due to decreased concentration of RBC
what does haemolysis interfere with?
determinations of colour interference (spectrophotometric assays) or chemical interactions
haematology
what is icterus?
yellow serum/plasma
what effect does jaundice have on biochemistry?
depends on the biochemistry being done
what is icterus/jaundice due to?
increased bilirubin - pathological
does jaundice interfere with haematology?
no
does icteric plasma/serum interfere with assays?
yes - should be rerun with a hopefully better sample
what are the main 5 types of in-house biochemistry analysers?
glucometres
‘dry’ biochemistry analysers (reagents are dry)
‘wet’ biochemistry analysers (reagents are wet)
electrolyte analyser
blood gas analyser
describe the basic principle of spectrophotometry
light source is passed through the sample and based on how much light of a particular wavelength is absorbed we can calculate concentration of the compound of interest
what law is spectrophotometry based on?
beers law
absorbance = constant + light pathway distance through cavette - concentration of substance of interest
what factors can interfere with spectrophotometry?
haemolysis and lipaemia can affect how much light is absorbed and can affect accuracy of results
how can haemolysis and lipaemia be prevented?
fasting and careful collection/handling
describe the best way to prevent haemolysis at all steps
careful sampling technique
avoid delays to sample processing
separate plasma/serum from sample as early as possible
refrigerate separated sample
what parameters are used to measure the function of the pancreas?
PLI and TLI
what does PLI stand for?
pancreatic lipase immunoreactivity
what does TLI stand for?
tripsenogen like immunoreactivity
what does elevated PLI indicate?
pancreatic injury
what does TLI indicate?
pancreatic functional mass - how much exocrine function remains (EPI)
what is EPI?
exocrine pancreatic insufficiency
why are the bile duct and pancreas so often linked in illness?
bile and pancreatic juice drain through a common duct. If there is cholestasis this can lead to secondary pancreatitis and vice versa
what 3 parameters are measured to show kidney function?
urea
creatinine
SDMA
what does increased urea in a biochemistry screen indicate?
dehydration, renal disease, urinary obstruction/rupture, heart disease
what does decreased urea in a biochemistry screen indicate?
liver failure as urea is made in the liver
what does increased creatinine in a biochemistry screen indicate?
as with urea: dehydration, renal disease, urinary obstruction/rupture, heart disease in medium and large dogs
what does decreased creatinine in a biochemistry screen indicate?
muscle wastage (very small dogs and cats)
is SDMA affected by muscle mass?
no, unlike creatinine
what must urea, creatinine and SDMA be assessed in conjunction with?
urine analysis
what do urea, creatinine and SDMA show?
glomerular filtration rate
how are urea, creatinine and SDMA linked?
tend to increase and decrease together
define azotemia
increased serum urea and/or creatinine concentration
where is azotemia detected?
in the blood
what are the 3 types of azotemia?
pre-renal
renal
post-renal
what can cause pre-renal azotemia?
decreased renal perfusion (hypovolaemia, dehydration)
what can cause renal azotemia?
disease - only seen when 60-70% of nephrons are damaged
what causes post-renal azotemia?
urinary tract obstruction or rupture
define uremia
specific clinical syndrome (vomiting, anorexia and ulcreation)
when does uremia develop?
when high levels of toxic substances (e.g. urea, creatinine)are present in the blood (azotemia) which causes reduced renal excretion
what is CK enzyme linked with?
muscle damage
what would increased CK indicate?
skeletal muscle damage or haemolysis
what other parameters may be increased alongside CK?
ALT and AST as they are also found within muscle
what is the totel proteins measurement formed of?
albumin + globulin
where is albumin produced?
liver
how can total proteins be measured?
refractometry
biochemistry analyser
what are the issues with refractometry when calculating total proteins?
may also measure glucose, cholesterol, urea etc, and is affected by lipaemia
what information about total protein will be given by biochemistry analyser?
albumin value and total protein
how can globulin be calculated from biochemistry analyser results?
total protein - albumin = globulin
what is the role in the blood of proteins?
maintaining oncotic pressure as well as individual protein functions
how may age affect total proteins?
increased in neonates after colostrum ingestion, decreases as maternal antibodies drop
what is the most common protein in the blood?
albumin
what can cause increased albumin in the blood?
dehydration
what can cause reduced albumin in the blood?
mild inflammation liver disease kidney disease GI disease haemorrhage
what is SPE?
serum protein electrophoresis
what is SPE used for?
evaluating the different groups of protein in serum
what do protein fractions migrate according to, during SPE?
size and charge
what can cause high glucose?
diabetes
stress
steroid therapy
what can cause low glucose?
wrong blood tube/time delay insulinoma insulin overdose some tumors (very metabolically active)
what does fructosamine indicate?
glucose level for past 2-3 weeks
what is fructosamine used to diagnose?
diabetes
what are the common liver parameters measured during biochemistry analysis?
ALT, GLDH, ALP, GGT, bilirubin, bile acids
what are the common kidney parameters measured during biochemistry analysis?
urea, creatinine, SDMA and urine analysis
what are the common pancreas parameters measured during biochemistry analysis?
PLI, TLI
what are the common muscle parameters measured during biochemistry analysis?
CK, AST
what are the common protein parameters measured during biochemistry analysis?
TP, albumin, APPs (acute phase proteins)
what are the common electrolyte parameters measured during biochemistry analysis?
Na, K, Cl, Ca, PO4
what are the common glucose parameters measured during biochemistry analysis?
fructosamine
what are the common lipid parameters measured during biochemistry analysis?
triglycerides and cholesterol
what are the common hormone parameters measured during biochemistry analysis?
T4, cortisol, TSH, ACTH, insulin
what are the common haematology parameters measured during biochemistry analysis?
RBC, WBC, Plt, blood smear
what are the common haemostasis parameters measured during biochemistry analysis?
PT, aPTT, TCT, BMBT
what can cause increased total proteins?
false high with lipaemia
dehydration
inflammation
neoplasia
what can cause decreased total proteins?
haemorrhage GI disease very young animals renal disease hepatic disease
which conditions have the largest effect on albumin?
renal disease
hepatic disease
what is haemostasis?
the ability to stop bleeding
what does defective haemostasis lead to?
leads to haemorrhage, generally known as coagulopathies
what does excessive haemostasis lead to?
leads to thrombotic events
what anticoagulant is used for clotting tests?
EDTA
what are the most important considerations when performing clotting tests?
respect amount of blood required
perform clean venapuncture
separate plasma ASAP
freeze if sending away
what are the 3 stages of haemostasis?
primary
secondary
tertiary
when does primary haemostasis occur?
first stage, occurs when the needle punctures the vein and lasts seconds to minutes
what is primary haemostasis associated with?
platelet numbers, function and vascular function leading to the formation of the platelet plug
what does primary haemostasis require?
enough platelets
platelets to function properly
von Wilburn factor
what happens during secondary haemostasis?
stabilisation of clot in minutes to hours
what is secondary haemostasis associated with?
formation of fibrin mesh which stabilises platelet plug
what happens during tertiary haemostasis?
return to normal vascular flow
what is tertiary haemostasis associated with?
breakdown of the clot
what can be used to measure tertiary haemostasis?
D-Diamers
what are the requirements for normal primary haemostasis?
vessel wall
platelets in correct number and functionin gproperly
vWF
what are the tests available for primary haemostasis?
BMBT
platelet count
platelet function testing
histopathology
what are the common disorders of primary haemostasis?
IMTP - immune mediated thrombocytopenia
vW disease
angiostrongylus
what are the requirements for normal secondary haemostasis?
coagulation factors
calcium
vitamin K
fibrin
what are the tests available for secondary haemostasis?
Activated Clotting Time
Prothrombin Time
Activated Partial Thromboplastin Time
Thrombin Clotting Time
what are the common disorders of secondary haemostasis?
congenital factor deficiencies
warfarin toxicity
severe liver disease
angiostrongylus
what are the requirements for normal tertiary haemostasis?
tissue plasminogen activator (tPA)
plasmin
what are the tests available for tertiary haemostasis?
FDPs
D-Diamers
what are the common disorders of tertiary haemostasis?
DIC
What is BMBT?
buccal mucosal bleeding time
how is BMBT tested?
under sedation - standardised cut made on patients gum. Blood collected with filter paper, taking care not to disturb the clot, and is absorbed until it stops
what are the normal BMBT for cats and dogs?
normal dog <3.3 mins
GA dog <4 mins
GA cat <3.3 mins
what causes prolonged BMBT?
moderate to severe thrombocytopenia
thrombopathia - inherited or acquired
vWM disease
what screening can BMBT be used for?
pre op of breeds at risk of vWD e.g. doberman
how quickly will hemorrhage start after trauma in a patient with primary haemostasis disorders?
immediately
what type of bleeding is typically seen in a patient with primary haemostasis disorders?
capillary bleeding: skin, mucosa, epistaxis, GI and urinary tract.
petechail haemorrhage
ecchymosis (bruising)
how quickly will hemorrhage start after trauma in a patient with secondary haemostasis disorders?
usually a small delay
what type of bleeding is typically seen in a patient with secondary haemostasis disorders?
internal bleeding: joints, muscles, body cavity (pleural, mediasteinum)
haematomas
haemoartroses
what bleeding is typically seen in a patient with both primary and secondary haemostasis disorders?
GI bleeding meleana haematoemesis haematochezia epistaxis haematauria
what is thrombocytopenia?
low platelets
how is activated clotting time tested?
whole blood is collected and allowed to clot in commercially available tubes
what are the normal activated clotting time values for dogs?
<90 sec
what are the normal activated clotting time values for cats?
<60 sec
when is activated clotting time prolonged?
if there is a marked deficiency in one/several clotting factors
affected by platelet number and function
what does the clotting cascade result in?
fibrin (clot)
what are the 2 branches of the clotting cascade
extrinsic
intrinsic
what can be used to measure extrinsic pathway of the clotting cascade?
prothrombolin time (PT)
what can be used to measure intrinsic pathway of the clotting cascade?
activated partial thromboplastin time (APTT)
what are PT and APTT tests checking for?
presence of clotting factors and function
what causes prolonged PT and APTT tests?
Increase in clotting times due to reduction in clotting factor (if individual factor is reduced to <20-30% of normal concentration)
where are most clotting factor and cofactors made?
in the liver
where do most clotting factors and cofactors circulate?
in the blood
what can be used to test for fibrinolysis?
FDPs
D Diamers
why to FDPs elevate?
increase on clot formation and lysis as well as inflammation
why do D Diamers elevate?
increases with clot formation and lysis
what are D Diamers?
degradation products of cross linked fibrin (stable clot)
when may elevated D Diamers be seen?
post surgery/trauma
DIC secondary to inflammation
neoplasia
thromboembolytic disease
