MAACB Prep Flashcards
(102 cards)
1
Q
Name 3 types of potentiometric electrode
A
1- redox
2- ion selective membrane ( glass and polymer)
3- PCO2
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
3
Q
Name 3 enzymes that can be used to measure glucose
A
Hexokinase
Glucose dehydrogenase
Glucose oxidase
4
Q
What is the most widely used glucose method with laboratories
A
Hexokinase coupled with glucose dehydrogenase , measuring NADH
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
6
Q
How many umol of glucose is consumed for every umol NADH?
A
1
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
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%
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
10
Q
What is the linearity of glucose HK Abbott method
A
0.28 - 44.40mmol/L
11
Q
What kind of interference does haemolysis cause Glucose HK spectro method ?
A
Haem concentrations >5g/L causes negative interference
12
Q
What kind of interference does ict and lip cause glucose HK method ?
A
Positive interference >55mmol/L
13
Q
Fasting glucose reference range ?
A
Adult- 2.8-5.6 mol/L
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
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
16
Q
Where is albumin synthesised ?
A
Parenchymal cells of the liver
+ fall in oncotic pressure
- IL6
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
18
Q
Possible causes of low albumin by decreased synthesis?
A
Inadequate nitrogen intake
Malabsorption
Chronic liver disease
19
Q
Possible cause of low albumin by increased catabolism?
A
Sepsis
20
Q
Possible cause of low albumin by redistribution?
A
Ascites
Oedema
Sepsis
21
Q
Possible cause of low albumin by increased loss?
A
Protein-losing enteropathy
Nephrotic syndrome
Loss of plasma eg.burns
22
Q
How do you convert mg/dl to mol/L?
A
Mol/L= (mg/dL X 10) / Mw
23
Q
List 4 hormones increased by stress
A
Glucocorticoids eg cortisol
Catecholamines eg epinephrine
Growth hormone
Prolactin
24
Q
List causes of elevated urate
A
Gout Renal failure Leukaemia Multiple myeloma Toxiemia of pregnancy
25
List causes of raises bilirubin
Increased hepatocellular damage
Infra and extra hepatic biliary tree obstruction
Haemolytic disease
Neonatal physiological jaundice
26
List causes of in vitro haemolysis
Incorrect needle size
Incorrect tube size selection (difficult bleed)
Prolonged tourniquet
Vigorous mixing of tubes
27
List biochemical analytes that change with dehydration
```
Urea
Creatinine
Sodium
HB
Protein
ADH
```
28
What is the ref method for lactate ?
There isn’t one
29
What is the reaction for the lactate oxidase method?
L-lactate + O2 —-LOD—-> Pyruvate + H2O2
H2O2 + 4-AA+ H+ ——Peroxidase—> chromogen + 2H2O
548nm
Fluoride oxalate sample
30
What is precision?
Closeness of agreement between independent results obtained under stipulated conditions.
31
What is repeatability ?
Closeness of agreement between results of successive measurements under the same conditions (within run precision )
32
What is reproducibility?
Closeness of agreement between results of measurements performed under changes conditions ( time , operators , calibrators )
33
What is drift?
Instrument or reagent instability over time
34
What is accuracy?
Closeness of the agreement between the result of a measurement and the true concentration of an analyte.
35
What is the limit of blank?
Highest apparent concentration in analyte-free material. 20 rots of sample with no analyte (0 calibrator)
LOB= mean(blank) + 1.645(SD blank)
36
What is LOD?
Lowest concentration confidently separated from blank 20 rots if verifying /60 if establishing)
LOD = LOB + 1.64( SD low conc sample)
37
What is LOQ?
Aka functional sensitivity-limit of reporting . Lowest concentration with acceptable reproducibility CV <20%
38
What is analytical sensitivity?
The smallest quantities that the assay measures
| LOB, LOD, LOQ
39
Define positive likelihood ratio, how is it calculated and how does it influence clinical decision making?
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
40
Name 4 methods for urinary free cortisol
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
41
What do the corticotroph cells of the anterior pituitary gland synthesize and secrete and to which receptor do they bind?
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
What are the series of reactions used for alkaline phosphatase measurement ?
4-nitrophenylphosphate(colourless) + H2O ——Alp , Mg2+—> 4-nitrophenoxide (benzenoid form) + HPO4 ——pH10.3–> 4-nitrophenoxide( yellow , quinod form)
43
List causes of an elevated serum cholesterol
```
Familial hypercholesterolemia
Cushing’s syndrome
Hypothyroidism
Pregnancy
2ndary to obstructive liver disease
```
44
List sources of alkaline phosphatase and how to differentiate them.
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
List causes of hypovolaemic hyponatraemia with low serum osmolality
Urine Na >20 & UOsm> 450 = renal loss: Addisons , salt losing nephritis, cerebral salt loss
Urine Ana <20 & UOsm >450 = GIT loss, sweat
46
List causes of euvolaemic hyponatraemia with low serum Osm
UNa >20 & UOsm >100 = SIADH, hypothyroidism, cortisol def
UNa variable & UOsm <100 = severe polydipsia , beer potomania, excess IV fluids
47
List causes of hypervolaemic hyponatraemia (low serum osmolality)
UNa>20 ARF of CRF
| UNa <20 nephrotic cirrhosis CCF
48
After a water deprivation test what urine osmolality would indicate central (neurogenic ) diabetes insipidis?
<300 after fluid deprivation
| >800 after desmopressin
49
After water deprivatation test what urine osmolality would indicate nephrogenic DI
After Fluid deprivation UOsm <300
| After desmopressin UOsm <300
50
What UOsm results after water deprivation test would indicate primary polydipsia ?
After Fluid deprivation UOsm >800
| After desmopressin UOsm >800
51
After water deprivation test what UOsm results would indicate partial DI or polydipsia?
After Fluid deprivation UOsm 300-800
| After desmopressin UOsm <800
52
What is functionally sensitivity?
Lowest concentration of analyte that can be measured at a specified precision
53
What is a heterogeneous immunoassay?
Immunoassay formate that uses 2 phases , usually liquid and solid to separate reacted from unreacted components
54
What is a heterophilic antibody?
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
What is high dose hook effect?
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
What is prozone phenomenon ?
Apparently lower reactivity or nonreactivity caused by a relative antigen excess
57
What is osmolality ?
The measurement of the number of moles of particles per kilogram of water
58
What is osmolarity?
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
How do you calculate osmolarity(mOsm/L)?
Osmolarity(mOsm/L) = 2X Na + glucose + urea
60
Name 4 measurements that depend on colligative properties
Colloid osmotic pressure(oncotic pressure)
Boiling point elevation
Freezing point depression
Vapor-pressure depression
61
Why is boiling point elevation measurement not used to determine osmolality?
Because proteins will coagulate, causing gross changes in the sample composition
62
What kind of thermometer is used to measure freezing point depression ?
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
Describe how freezing point depression is measured.
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
What is the osmolar gap?
Serum osmolality gap= measured - calculated
| The average gap is near zero
65
What is the ideal osmolality gap for mannitol therapy?
Mannitol is used to treat patients with Edema , especially cerebral edema.
Aim for osmolal gap of 10 - 50 mOsm/L
66
How do you calculate stool osmolal gap
Stool osmolal gap= measured osmolality (stool) - 2([Na+] + [K+]) stool
67
Why is stool osmolal gap calculated?
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
What is vitamin D2 also known as where is it obtained?
Ergocalciferol
Obtained through diet or supplements
Metabolises to 25(OH) vitaminD2
69
What is vitamin D3 also known as and where is it obtained?
Cholecalciferol
Mainly produced in skin through UV B exposure
Metabolised to 25(OH) vitamin D3
70
What are some advantages / challenges of using the enzymatic method for HbA1c
Advantages: no analytical interference from sub variants
Challenge: unable to detect Hb variants
71
What is the principle of the enzymatic HbA1c method?
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
What is the principle of the immunoassay for HbA1c?
Uses Abs which target the first 4-10a.a at the N-terminal of the beta chain
73
List advantage/challenges of using immunoassay for HbA1c
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
What is the Principle of boron are affinity HPLC for HbA1c?
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
Name advantage/challenges of boron are affinity HPLC for HbA1c
Advantages: considered to have the least analytical interference from Hb variants.
Challenge: measures all glycated Hb and cannot detect Hb variants
76
What is the principle of ion-exchange HPLC for HbA1c measurement?
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
Name advantages/challenges to using ion-exchange HPLC for HbA1c measurement
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
What is the principle of capillary electrophoresis for HbA1c testing?
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
List advantages/challenges to CE for HbA1c
Advantages: high chromatic resolution and resulting ability to detect many Hb variants.
Challenge: throughput
80
List four methods of HbA1c measurement
```
Enzymatic
Immunoassay
Capillary electrophoresis
HPLC- ion- exchange
HPLC- boronate affinity
```
81
Define diagnostic sensitivity
How good a test is at identifying those with disease. True positives
Sensitivity = TP/TP + FN
82
Define diagnostic specificity
How good at identifying those without disease. True negatives
Spec= TN/TN+FP
83
List causes of in Vitro haemolysis
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
Name reasons for in vivio haemolysis
Hereditary - sickle cell
Acquired (auto immune)
Transfusion reactions , heart valves
85
What is the reference method for measurement of haemolysis?
Cyanomethaemoglobin
86
List the mechanisms of haemolysis interference
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
List mechanisms of interference for lipids
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
What can give a falsely high lipaemic index?
IgM paraproteins
89
What are the mechanisms of interference for Icteria?
Spectral - interference with Jaffe creatinine method
| Chemical - can react with H2O2 - chol, trig, urate, Fe and enzymatic creatinine method
90
What can be added to remove bilirubin (ictera ) interference?
Ferrycyanide solution destroys bilirubin.
| Or alternative method eg chromatography
91
What areas need to be checked when testing for carryover?
Probe wash process
Curette wash process
Tube components
92
How do heterophile antibodies give false positives in sandwich immunoassays ?
By forming a bridge between two antibodies as the analyte would.
93
What is a HAMA?
Human anti-mouse antibody
94
Name a protein that can behave like a heterophile antibody
Rheumatoid factor
95
At what protein levels should a direct ISE method be used for na
<40 (pseudo hyper state)
| >100g/L ( pseudo hypo state)
96
What is Rayleigh scattering?
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
What is Raleigh-Debye scattering?
When particle size is only a little smaller than , or is equal to the wavelength , asymmetric forward scattering .eg IgM ,chylomicrons
98
What is Mie scattering?
Asymmetric forward scattering increases significantly when particle size is larger than the wavelength eg RBC, bacteria
99
Name examples of Atomic absorption spectrometry
Flame atomic absorption spectroscopy
| Graphite furnace atomic absorption spectroscopy
100
Name types of atomic emission spectrometry
Flame atomic emission spectroscopy
| ICP -OED ( inductively coupled plasma - optical emission spectroscopy
101
What kind of sample prep is required for ICPMS?
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
Name possible sources of error in ICPMS
```
Memory effects
Contamination
Matrix effects
Isobaric elements
Double charged ions
Polyatomic ions
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