MAACB Prep Flashcards

1
Q

Name 3 types of potentiometric electrode

A

1- redox
2- ion selective membrane ( glass and polymer)
3- PCO2

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2
Q

Define precision , repeatability and reproducibility

A

Precision is the closeness of agreement of independent results , measured under stipulated conditions
Repeatability is within run precision.
Reproducibility is under changed conditions eg time, operator , calibrators

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3
Q

Name 3 enzymes that can be used to measure glucose

A

Hexokinase
Glucose dehydrogenase
Glucose oxidase

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4
Q

What is the most widely used glucose method with laboratories

A

Hexokinase coupled with glucose dehydrogenase , measuring NADH

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5
Q

What is the equation for the glucose Hexokinase method

A

Glucose + MgATP —HK—->G6P + MgADP

G6P + NAD —G6PD—-> 6-phosophogluconolactone + NADH

NADH measured at 340nm

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6
Q

How many umol of glucose is consumed for every umol NADH?

A

1

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7
Q

How many calibration points and what calibrator is used for the Hexokinase glucose method

A

3 point cal

Blank plus MCC1 and MCC 2

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8
Q

What is the LOD, LOQ for serum glucose

A
LOD= 0.14mmol/L with 95%probability 
LOQ= 0.28mmol/L with CV 20%
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9
Q

What is the LOD and LOQ for urine/csf glucose

A
LOD= 0.056mmol/L 95%propbability
LOQ= 0.056mmol/L 20%CV
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10
Q

What is the linearity of glucose HK Abbott method

A

0.28 - 44.40mmol/L

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11
Q

What kind of interference does haemolysis cause Glucose HK spectro method ?

A

Haem concentrations >5g/L causes negative interference

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12
Q

What kind of interference does ict and lip cause glucose HK method ?

A

Positive interference >55mmol/L

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13
Q

Fasting glucose reference range ?

A

Adult- 2.8-5.6 mol/L

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14
Q

Describe the glucose oxidase reaction ?

A

Glucose + O2 —GO—-> gluconlactone + H2O2

H2O2 + chromogenic oxygen acceptor (e.g o-diasidine ) —peroxidase—> chromogen colour + H2O

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15
Q

What is mutarotase used for?

A

In kits to convert alpha D-glucose to beta D-glucose for use in glucose oxidase polarography reaction - measuring oxygen consumption

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16
Q

Where is albumin synthesised ?

A

Parenchymal cells of the liver
+ fall in oncotic pressure
- IL6

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17
Q

What is the function of albumin

A

Non-specific transport protein eg FFA, ca2+,Zn2+, unconjugated bilirubin and many drugs.
Contributes to oncotic pressure
Minor buffer of H+ ions

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18
Q

Possible causes of low albumin by decreased synthesis?

A

Inadequate nitrogen intake
Malabsorption
Chronic liver disease

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19
Q

Possible cause of low albumin by increased catabolism?

A

Sepsis

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20
Q

Possible cause of low albumin by redistribution?

A

Ascites
Oedema
Sepsis

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21
Q

Possible cause of low albumin by increased loss?

A

Protein-losing enteropathy
Nephrotic syndrome
Loss of plasma eg.burns

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22
Q

How do you convert mg/dl to mol/L?

A

Mol/L= (mg/dL X 10) / Mw

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23
Q

List 4 hormones increased by stress

A

Glucocorticoids eg cortisol
Catecholamines eg epinephrine
Growth hormone
Prolactin

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24
Q

List causes of elevated urate

A
Gout
Renal failure
Leukaemia 
Multiple myeloma 
Toxiemia of pregnancy
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25
Q

List causes of raises bilirubin

A

Increased hepatocellular damage
Infra and extra hepatic biliary tree obstruction
Haemolytic disease
Neonatal physiological jaundice

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26
Q

List causes of in vitro haemolysis

A

Incorrect needle size
Incorrect tube size selection (difficult bleed)
Prolonged tourniquet
Vigorous mixing of tubes

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27
Q

List biochemical analytes that change with dehydration

A
Urea
Creatinine
Sodium
HB
Protein
ADH
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28
Q

What is the ref method for lactate ?

A

There isn’t one

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29
Q

What is the reaction for the lactate oxidase method?

A

L-lactate + O2 —-LOD—-> Pyruvate + H2O2
H2O2 + 4-AA+ H+ ——Peroxidase—> chromogen + 2H2O
548nm
Fluoride oxalate sample

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30
Q

What is precision?

A

Closeness of agreement between independent results obtained under stipulated conditions.

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31
Q

What is repeatability ?

A

Closeness of agreement between results of successive measurements under the same conditions (within run precision )

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32
Q

What is reproducibility?

A

Closeness of agreement between results of measurements performed under changes conditions ( time , operators , calibrators )

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33
Q

What is drift?

A

Instrument or reagent instability over time

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34
Q

What is accuracy?

A

Closeness of the agreement between the result of a measurement and the true concentration of an analyte.

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35
Q

What is the limit of blank?

A

Highest apparent concentration in analyte-free material. 20 rots of sample with no analyte (0 calibrator)

LOB= mean(blank) + 1.645(SD blank)

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36
Q

What is LOD?

A

Lowest concentration confidently separated from blank 20 rots if verifying /60 if establishing)

LOD = LOB + 1.64( SD low conc sample)

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37
Q

What is LOQ?

A

Aka functional sensitivity-limit of reporting . Lowest concentration with acceptable reproducibility CV <20%

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38
Q

What is analytical sensitivity?

A

The smallest quantities that the assay measures

LOB, LOD, LOQ

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39
Q

Define positive likelihood ratio, how is it calculated and how does it influence clinical decision making?

A

LR+ is the probability that a person with the disease test positive (TP) divided by the probability that a person without the disease test positive (FP)
LR+= sensitivity/1-specificity
LR- = 1-sensitivity/specificity
Applying likelihood ratios can change the probability of a diagnosis eg low enough to rule out disease or high enough to start treatment

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40
Q

Name 4 methods for urinary free cortisol

A

LC-MS/MS
Beckman coulter -EIA(comp chemilum) 30mins
Roche - ECLIA ( comp electro chemilum IA) 18mins NB:biotin
Abbott -CMIA( comp chemilum Microparticles IA) 30 mins

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41
Q

What do the corticotroph cells of the anterior pituitary gland synthesize and secrete and to which receptor do they bind?

A

ACTH
Binds to the melanocortin receptor 2. Needs presence of MC2 accessory protein to respond to ACTH.
ACTH acts through the formation of cAMP which facilitates the transfer of cholesterol into the mitochondrial inner membrane for the synthesis of adrenal steroids

42
Q

What are the series of reactions used for alkaline phosphatase measurement ?

A

4-nitrophenylphosphate(colourless) + H2O ——Alp , Mg2+—> 4-nitrophenoxide (benzenoid form) + HPO4 ——pH10.3–> 4-nitrophenoxide( yellow , quinod form)

43
Q

List causes of an elevated serum cholesterol

A
Familial hypercholesterolemia
Cushing’s syndrome 
Hypothyroidism
Pregnancy
2ndary to obstructive liver disease
44
Q

List sources of alkaline phosphatase and how to differentiate them.

A

Liver and bone chromosome 1
Intestinal & placental chromosome 2

Alkaline phosphatase isoenzyme electrophoresis - separates different isoforms
Placental is heat labile and may need to be treated to differentiate

45
Q

List causes of hypovolaemic hyponatraemia with low serum osmolality

A

Urine Na >20 & UOsm> 450 = renal loss: Addisons , salt losing nephritis, cerebral salt loss

Urine Ana <20 & UOsm >450 = GIT loss, sweat

46
Q

List causes of euvolaemic hyponatraemia with low serum Osm

A

UNa >20 & UOsm >100 = SIADH, hypothyroidism, cortisol def

UNa variable & UOsm <100 = severe polydipsia , beer potomania, excess IV fluids

47
Q

List causes of hypervolaemic hyponatraemia (low serum osmolality)

A

UNa>20 ARF of CRF

UNa <20 nephrotic cirrhosis CCF

48
Q

After a water deprivation test what urine osmolality would indicate central (neurogenic ) diabetes insipidis?

A

<300 after fluid deprivation

>800 after desmopressin

49
Q

After water deprivatation test what urine osmolality would indicate nephrogenic DI

A

After Fluid deprivation UOsm <300

After desmopressin UOsm <300

50
Q

What UOsm results after water deprivation test would indicate primary polydipsia ?

A

After Fluid deprivation UOsm >800

After desmopressin UOsm >800

51
Q

After water deprivation test what UOsm results would indicate partial DI or polydipsia?

A

After Fluid deprivation UOsm 300-800

After desmopressin UOsm <800

52
Q

What is functionally sensitivity?

A

Lowest concentration of analyte that can be measured at a specified precision

53
Q

What is a heterogeneous immunoassay?

A

Immunoassay formate that uses 2 phases , usually liquid and solid to separate reacted from unreacted components

54
Q

What is a heterophilic antibody?

A

Artefact seen in sandwich immunoassays, in which patients endogenous antibody to reagent antibodies links labeled antibody to capture antibody in the absence of antigen. Results in false elevations of analyte.

55
Q

What is high dose hook effect?

A

Artefact seen in sandwich immunoassay in which labeled antibody bound in the sandwich decreases in the presence of excess antigen. Results in falsely low assay results

56
Q

What is prozone phenomenon ?

A

Apparently lower reactivity or nonreactivity caused by a relative antigen excess

57
Q

What is osmolality ?

A

The measurement of the number of moles of particles per kilogram of water

58
Q

What is osmolarity?

A

An alternative measure of colligative property that is clinically equivalent to osmolality;
Measurement of the number of moles of particles per litre of water.

59
Q

How do you calculate osmolarity(mOsm/L)?

A

Osmolarity(mOsm/L) = 2X Na + glucose + urea

60
Q

Name 4 measurements that depend on colligative properties

A

Colloid osmotic pressure(oncotic pressure)
Boiling point elevation
Freezing point depression
Vapor-pressure depression

61
Q

Why is boiling point elevation measurement not used to determine osmolality?

A

Because proteins will coagulate, causing gross changes in the sample composition

62
Q

What kind of thermometer is used to measure freezing point depression ?

A

A thermistor ( thermal resistor) a semiconductor made from a mixture of oxides of transition metals such as manganese, cobalt, and nickel. Thermistors become better conductors as temperature rises. The conductance or resistance of metals can be related to the temperature and hence to the osmolality

63
Q

Describe how freezing point depression is measured.

A

1- sample cooled rapidly by cold block to -5’C
2- vigorous agitation induces the crystallisation process. Once crystals begin to form , additional water molecules are added rapidly to the ice crystals. Heat is released in the freezing process , this raises the Temperature of the sample until freezing stops and equilibrium temperature establishes.
This temp is inversely related to the osmolality of the sample

64
Q

What is the osmolar gap?

A

Serum osmolality gap= measured - calculated

The average gap is near zero

65
Q

What is the ideal osmolality gap for mannitol therapy?

A

Mannitol is used to treat patients with Edema , especially cerebral edema.
Aim for osmolal gap of 10 - 50 mOsm/L

66
Q

How do you calculate stool osmolal gap

A

Stool osmolal gap= measured osmolality (stool) - 2([Na+] + [K+]) stool

67
Q

Why is stool osmolal gap calculated?

A

To differentiate causes of chronic diarrhoea
Gap <50mOsm/L = secretion all diarrhoea
>50mOsm/L = Unabsorbed material eg food. Excessive use of laxatives

Measured osm should be similar to plasma . A hypo-osm< 280mOsm/L may be factitious diarrhoea (munchausen)

68
Q

What is vitamin D2 also known as where is it obtained?

A

Ergocalciferol
Obtained through diet or supplements
Metabolises to 25(OH) vitaminD2

69
Q

What is vitamin D3 also known as and where is it obtained?

A

Cholecalciferol
Mainly produced in skin through UV B exposure
Metabolised to 25(OH) vitamin D3

70
Q

What are some advantages / challenges of using the enzymatic method for HbA1c

A

Advantages: no analytical interference from sub variants
Challenge: unable to detect Hb variants

71
Q

What is the principle of the enzymatic HbA1c method?

A

Uses an enzyme which specifically cleaves the N-terminal valine.
Lysed RBCs undergo proteolytic digestion to release substrate fructosyl valine
Fructosyl valine + O2 —-FVO+ FADH2–> H2O2 + valine + glucosone
H2O2 + leuco dye —-HPO—> colorimetric result 670nm

72
Q

What is the principle of the immunoassay for HbA1c?

A

Uses Abs which target the first 4-10a.a at the N-terminal of the beta chain

73
Q

List advantage/challenges of using immunoassay for HbA1c

A

Advantages: no analytical interference from most common Hb variants using newer gen assays
Challenge: unable to detect Hb variants, still susceptible to rare Hb variants

74
Q

What is the Principle of boron are affinity HPLC for HbA1c?

A

The structural difference between HbA0 & HbA1c resulted from the presence of the glucose group constitutes the basis for this method. Glycohaemoglobin binds affinity resin while nonglycated Hb passes through the column. M-aminophenyboronic acid binds glucose moiety to affinity resin

75
Q

Name advantage/challenges of boron are affinity HPLC for HbA1c

A

Advantages: considered to have the least analytical interference from Hb variants.
Challenge: measures all glycated Hb and cannot detect Hb variants

76
Q

What is the principle of ion-exchange HPLC for HbA1c measurement?

A

Uses buffers of increasing ionic strength to separate HbA1c from other Hb species by differences in charge. Concentration of each fraction is quantified by calculating the area under each peak

77
Q

Name advantages/challenges to using ion-exchange HPLC for HbA1c measurement

A

Advantage :ability to detect most common Hb variants can give presumptive ID
Challenges: can have interference from Hb variant that co-elite with peaks of interest.
Schiff
Labour intensive
TAT

78
Q

What is the principle of capillary electrophoresis for HbA1c testing?

A

Separates by charge and mass. Charged molecules are highly resolved by electrophoretic mobility and their separation depends on electrolyte pH and electro-osmotic flow.

79
Q

List advantages/challenges to CE for HbA1c

A

Advantages: high chromatic resolution and resulting ability to detect many Hb variants.
Challenge: throughput

80
Q

List four methods of HbA1c measurement

A
Enzymatic 
Immunoassay 
Capillary electrophoresis 
HPLC- ion- exchange
HPLC- boronate affinity
81
Q

Define diagnostic sensitivity

A

How good a test is at identifying those with disease. True positives

Sensitivity = TP/TP + FN

82
Q

Define diagnostic specificity

A

How good at identifying those without disease. True negatives

Spec= TN/TN+FP

83
Q

List causes of in Vitro haemolysis

A

Collection into syringe-drawing up blood under pressure
Collection with small needle large tube -vacuum through small hole
Transfer of sample from syringe to tube
Alcohol contamination with capillary collection
Freezing sample

84
Q

Name reasons for in vivio haemolysis

A

Hereditary - sickle cell
Acquired (auto immune)
Transfusion reactions , heart valves

85
Q

What is the reference method for measurement of haemolysis?

A

Cyanomethaemoglobin

86
Q

List the mechanisms of haemolysis interference

A

Addiitive - released from cells( K, LD, AST, Phos, Fe, folate , ammonia)

Spectral - method wavelength around Hb peak (415, 540, 570nm) eg GGT, ALP

Chemical - cross -reactivity of cell analytes and method eg adenylte kinase in RBCs cross reacts with CK, proteolytic enzymes can destroy small peptides eg insulin, glucagon, calcitonin, PTH, ACTH and TnT

Dilutional - gross haemolysis (Na, Cl etc)

87
Q

List mechanisms of interference for lipids

A

Spectrophotometric - methods using lower wavelength
Volume displacement - indirect ISE methods dilute the sample enhancing solution effect
Physical - block Ag-Ab complex formation, bind lipophilic compounds eg drugs

88
Q

What can give a falsely high lipaemic index?

A

IgM paraproteins

89
Q

What are the mechanisms of interference for Icteria?

A

Spectral - interference with Jaffe creatinine method

Chemical - can react with H2O2 - chol, trig, urate, Fe and enzymatic creatinine method

90
Q

What can be added to remove bilirubin (ictera ) interference?

A

Ferrycyanide solution destroys bilirubin.

Or alternative method eg chromatography

91
Q

What areas need to be checked when testing for carryover?

A

Probe wash process
Curette wash process
Tube components

92
Q

How do heterophile antibodies give false positives in sandwich immunoassays ?

A

By forming a bridge between two antibodies as the analyte would.

93
Q

What is a HAMA?

A

Human anti-mouse antibody

94
Q

Name a protein that can behave like a heterophile antibody

A

Rheumatoid factor

95
Q

At what protein levels should a direct ISE method be used for na

A

<40 (pseudo hyper state)

>100g/L ( pseudo hypo state)

96
Q

What is Rayleigh scattering?

A

When particle size is significantly smaller than the wavelength. Light is scattered symmetrically in the direction of the incoming light as well as against incoming light.

97
Q

What is Raleigh-Debye scattering?

A

When particle size is only a little smaller than , or is equal to the wavelength , asymmetric forward scattering .eg IgM ,chylomicrons

98
Q

What is Mie scattering?

A

Asymmetric forward scattering increases significantly when particle size is larger than the wavelength eg RBC, bacteria

99
Q

Name examples of Atomic absorption spectrometry

A

Flame atomic absorption spectroscopy

Graphite furnace atomic absorption spectroscopy

100
Q

Name types of atomic emission spectrometry

A

Flame atomic emission spectroscopy

ICP -OED ( inductively coupled plasma - optical emission spectroscopy

101
Q

What kind of sample prep is required for ICPMS?

A

Blood , urine or plasma - dilute and shoot. Acid -2% nitric acid, base - 1% ammonium , 0.1% EDTA, 0.05% triton x-100
Tissue sample , hair, nail - chemical digestion - concentrated acid HCL, or base TMAH

102
Q

Name possible sources of error in ICPMS

A
Memory effects
Contamination 
Matrix effects 
Isobaric elements
Double charged ions
Polyatomic ions