LEC LIPIDS PART 2 - LIPID DETERMINATION Flashcards
(183 cards)
1
Q
Disorders of lipids
A
Arteriosclerosis, obesity, hypertension, DIABETES MELLITUS and other abnormalities
2
Q
Good prognosis if lipid
A
Early detection of deranged blood lipid profile
3
Q
age for screening for lipid
A
20 years old, if the test is normal repeat again after 5 yrs
if not, depends on the physician
4
Q
- Indications for Lipid Profile include:
A
- ⁃ Screening for primary & secondary
hyperlipidemias - ⁃ Monitoring for risk of atherosclerosis
3.⁃ Monitoring treatment of
hyperlipidemias/Dyslipidemia
5
Q
preanalytical considerations
biological variations such as
A
- Age: cholesterol levels increase with age.
- Sex: women have lower level than men except in childhood & after early 50’s.
- Season: cholesterol levels are slightly higher in cold periods.
- Food intake: daily intake of fat increases cholesterol levels.
- Medical conditions: thyroid, liver, and kidney diseases
- Acute illness
- Life-style: higher in sedentary and poor diet habits
6
Q
explain the cholesterol level in terms of age
A
it increases with age
7
Q
describe the gender relation of cholesterol
A
women have lower level than men except in childhood and after early 50`s
high in menopausal stage
8
Q
can season affect cholesterol?
A
higher in cold cholesterol
9
Q
why do menopausal women has high chole or high lipid profile?
A
because estrogen came from chole, if the cholesterol are can’t be converted into estrogen any longer, it will be deposited and stuck in the blood stream elevating lipids in the body
10
Q
daily intake of fat increases cholesterol levels
true or false
A
true
11
Q
in hyperthyroidism, chole is low because?
A
because of fast metabolism
12
Q
if we have liver disease, do we have high or low chole?
A
low, 85% of chole is produced by the liver. If the liver is defected, there will be no organ that will produce it
13
Q
describe the lipid profile of chronic alcoholic patients
A
low
14
Q
describe the lipid profile of moderate alcoholic patients
A
high
15
Q
components that clears the chylomicron
A
lipoprotein lipase that are activated by apo C2
16
Q
Patient should fast for ___hours before sampling
A
12
17
Q
Chylomicrons are cleared within__hrs and their presence
after 12hrs fast is abnormal.
A
6-9
18
Q
Posture: Decreases of as much_________ of TC, LDL-C, HDL-C and apo-A-1 and B
A
10 %
19
Q
correct position of getting sample
A
supine and lying position
20
Q
Patient to be seated for __ prior to sampling to prevent hemoconcentration.
A
5min
21
Q
Prolonged venous occlusion leads to increase in cholesterol conc by _______.
venous occlusion means tourniquet
A
10-15%
22
Q
Exercise: Mild exercise produces a slight decrease in
conc of _______________.
A
chole and triglycerides
23
Q
Those who walk for about 4 hours each week have an average cholesterol ___ lower and HDL-C ___ higher than inactive persons
A
5%; 3.4%
24
Q
lipids that has higher concentration in the midcycle of menstruation
A
plasma Cholesterol and Triglycerides
25
A high fat diet increases serum __
triglyceride
26
-Ingestion of monounsaturated fat reduces __.
cholesterol
27
__ conc is reduced when sucrose intake is reduced.
Plasma triglyceride
28
A high carbohydrate diet decreases the serum conc. of___
VLDL-C, TG,
cholesterol and protein.
29
Large protein meals at lunch or in the evening also ___ the serum
cholesterol for atleast 1 hour after a meal.
increase
30
In vegeterian individuals, conc. of LDL & VLDL-C are reduced by ___ and ____
LDL 37%
VLDL 12%
31
Smoking:
The plasma cholesterol, triglyceride and LDL cholesterol conc. are higher by about ___respectively in smokers
than in non smokers
3 %, 9.1 % and 1.7 %
cholesterol, 3 %,
triglyceride 9.1 %
LDL cholesterol 1.7 %
32
__ cholesterol is lower in smokers than in non smokers
HDL
33
When MODERATE amount of alcohol is ingested
for 1wk, the serum TG conc is increased by more than _________.
20 mg/dl
33
Prolonged moderate ingestion of alcohol may increase HDL-C conc
it will reduced the plasma concentration of
cholesterol ester transfer protein(CETP).
33
what sample can we use for lipid profile?
either plasma or serum
34
___ is preferred when lipoprotein are measured by ultracentrifugation and electrophoretic methods
Plasma
35
if plasma is used for lipid profile, what anticoagulant is the best choice?
edta
36
2 methods to detect lipoprotein using plasma
ultracentrifugation and electrophoretic methods
37
reference method for lipoprotein determination
ultracentrifugation
38
the only lipoprotein that is only in the origin during the electrophoresis
chylomicron
39
which sample tend to have lower concentration of lipids
venous or capillary?
and why?
capillary - same with glucose, because of water
40
an anticoagulant that will exert large osmotic effect
citrate
41
Citrate: exert large osmotic effect resulting in falsely __ plasma lipid and
lipoprotein concentration.
low
42
an anticoagulant that can alter electrophoretic mobility of
lipoproteins.
heparin
43
heparin can alter electrophoretic mobility of
lipoproteins because of its __
high molecular weight
44
electrophoresis is separation of components depending on the
sizes
45
EDTA plasma
are ___% lower than in serum.
3%
46
in terms of storing
TC, TG, HDL-C can be satisfactorily analyzed in ___ samples.
frozen
47
aside from total chole, hdl, and tg
-__ can also be measured in frozen samples.
Apolipoproteins
48
Cholesterol Estimation has 2 methods, what are they
chemical and enzymatic method
49
if we have a given serum and we need to convert it to plasma, what formula we should use?
serum x 3%
50
if we have plasma and we need to find the serum, what's the formula
X 1.03
51
we store sample or plasma in what temperature for a long time
- 70*
52
we store sample or plasma in what temperature for a short time (1-2 months)
-2 *
53
what is the former reference method for chemical estimation
abell kendall method
54
3 steps of abell kendall method
1. Cholesterol is HYDROLYZED with alcoholic KOH
2. Unesterified cholesterol is EXTRACTED with petroleum jelly/Hexane
3. Measured using the L-B Reaction (COLORIMETRY)
55
RESULT OF HYDROLYZING CHOLESTEROL
cholesterol ester AND FATTY ACIDS
removing cholesterol from fatty acids is
free cholesterols
56
how Liebermann-burchardt reaction works?
the unsterified extracted will combined with sulfuric acid and acetic anhydride to form a bluish green solution
unsterified Cholesterol + Sulfuric acid + Acetic anhydride bluish green solution
57
end product of liebermann-burchardt's reaction
bluish green solution called Cholestadienyl monosulfonic acid
58
purpose of acetic anhydride and sulfuric acid
color developers
59
another chemical method for cholesterol estimation that only uses 2 steps
Bloors Method
60
Bloors Method: - Principle: 2 step
1. Cholesterol is extracted using an alcohol ether mixture
2. Measured using the L-B Reaction
61
in bloor's method, the cholesterol is extracted using
alcohol ether mixture
62
in abell kendall method - the cholesterol is extracted using
petroleum jelly and hexane
63
in abell kendall method - the cholesterol is hydrolyzed using
alcoholic potassium hydroxide KOH
64
what is the principle of abell kendall method
3 step method - hydrolysis, extraction, colorimetry
65
what is the principle of enzymatic method
cholesterol oxidase method
66
Routine Lab - Assay of Choice for cholesterol
Cholesterol Oxidase Method
67
Cholesterol Oxidase Method is only specific for __
cholesterol esters
68
cholesterol esters are those cholesterols with ___
fatty acids
69
the cholesterol ester are hydrolyze by what enzyme to form free chole and fatty acids
cholesterol esterase
70
free cholesterol will be oxidized by what enzyme
cholesterol oxidase
71
free cholesterol will be oxidized by cholesterol oxidase to form
4 cholestene - 3 - one + h202
72
what is the goal of oxidizing free cholesterol in cholesterol oxidase method
to find the H202 which will be used in trinder's reaction
73
in trinders reaction we uses what enzyme
PEROXIDASE
74
what specific peroxidase is used for trinder's reaction
horse raddish peroxidase
75
what dye is used to form quinoneimine dye in trinder's reaction
4-aminophenazone
76
a sequential reaction in trinder's reaction
low chole shows
high chole show
low chole shows pink
high chole show red
77
in trinder's reaction or oxidase method, how many enzymes are used
3 - esterase , oxidase, peroxidase
78
cholesterol oxidation or trinder's method is read at what wavelength?
520 nm
79
cholesterol oxidase method has a linearity up to
600 - 700mg/dL (15.54 - 18.13mmol/L)
80
normal value of cholesterol
81
ADVANTAGE of enzymatic method
Precise and accurate
Lesser interferences -bilirubin, ascorbic acid, Hb
Smaller sample quantity
Rapid; does not require preliminary extraction step
Can be used to measure unesterified cholesterol by omitting de-esterification step
Mild reagents; better suited for automated analyzers
82
DISADVANTAGE of enzymatic reaction
They are not absolutely specific for
cholesterol.
Cholesterol oxidase reacts with other sterols e.g plant sterol
Ascorbic acid and Bilirubin interfere by consuming H202
Bilirubin interference can produce falsely high or low values
Significant only at conc >5mg/dL decreasing Cholesterol values by 5 -15%
83
INCREASED CHOLESTEROL are found in
1. Biliary cirrhosis
2. Hyperlipoproteinemia types
II, III, V
3. Nephrotic syndrome
4. Uncontrolled DM
5. Alcoholism
6. Primary Hypothyroidism
84
DECREASED CHOLESTEROL
1. Severe hepatocellular
disease (alcoholic liver
disease)
2. Malnutrition
3. Severe burns
4. Hyperthyroidism
5. Malabsorption syndrome
85
limit of interferences of bilirubin for chole
not greater than 200 mg/dl
86
limit of interferences of hemoglobin for chole
not greater than 20 mg/dl
87
more than 5 mg/dl in bilirubin will interfere to chole increasing the value of _- %
5-15%
88
the reference method for cholesterol
gas chromatography/mass spectrometry
gc ms
89
Cholesterol Desirable level:
: < 200mg/dL (< 5.2mmol/L);
90
in a modified abell kendall method, they use hexane, chloroform, and silisic acid
what is the purpose of chloroform in the process?
remove phospholipid
91
the color of the end product of the modified abell kendall method
pink
91
what kind of lipid is affected in a fasting specimen?
LDL and Triglyceride
91
in a modified abell kendall method, they use hexane, chloroform, and silisic acid
what is the purpose of silisic acid?
adsorption of phospholipid via chromatography
91
a post prandial lipemia
chylomicron
91
in non fasting sample, what type of triglyceride carrying lipoprotein is found
chylomicron
91
we use plasma - the preferred sample for measuring lipoprotein.
In ultracentrifugation, what substance do we use __ to separate lipoproteins based on density
potassium bromide
91
what apoprotein that can inhibit the function of apo c2?
apo protein c3
91
if we go under fasting, what type of triglyceride carrying lipoprotein can we observed on greatest amount?
vldl
91
what is the specific
1.063`
91
polyanions
heparin sulfate, dextran sulfate and phosphotungstate
91
divalent cations
Mg, Ca and Mn
92
Most commonly for HDL and is reasonably specific
POLYANION PRECIPITATION:
92
the cdc reference method
abell kendall
92
what's in it inside the cholesterol making them possible to be measured using colorimetric assay/chemical method
the presence of double bonds and hydroxyl groups of the esterols
92
what is the end product of the salkowski method
cholestadienyl disulfonic acid - red solution
92
Precipitating reagents such as ___ and ____ are used to remove all lipoproteins except
HDL
divalent cations and polyanions
93
how can ascorbic acid affect the result of cholesterol determination
falsely decrease
94
how can hemoglobin and bilirubin affects the result of cholesterol determination
falsely increase
95
how can high level of triglyceride affect hdl determination
elevated triglyceride results to incomplete sedimentation which is as well will cause the hdl result elevated
96
Similar to the HDL-C precipitation method but uses a
precipitant that is complexed to magnetic particle
MAGNETIC METHOD
97
This sediments and does not require centrifugation
MAGNETIC METHOD
98
Has been adapted for use in automated clinical chemistry
analyzers
MAGNETIC METHOD
99
It allows the supernatant to be analyzed without the
need to remove it from the sedimented complex
MAGNETIC METHOD
100
homogenous assay or direct hdl c assay is a type of what method
enzymatic method
101
HOMOGENOUS ASSAY (Direct HDL-C Assay):
Enzymatic method: First reagent - "___" non-HDLs
blocks
102
blocker for homogenous assay
Use of Antibodies or Polymers or complexing agent
e.g Cyclodextrin
103
HOMOGENOUS ASSAY (Direct HDL-C Assay) steps
Enzymatic method: First reagent - "blocks" non-HDLs
Use of Antibodies or Polymers or complexing agent
e.gCyclodextrin
Modification of cholesterol esterase and oxidase enzymes which makes them selective for HDL-C
Use of blanking step that selectively consumes cholesterol from non-HDL species
104
Reference method for HDL-C
estimation is
Three-step Procedure
105
comment about three step procedure
tedious and expensive
106
2 types of ULTRACENTRIFUGATION
Preparative Ultracentrifugation
Density gradient method
107
a type of centrifugation that Uses sequential density adjustments of serum to FRACTIONATE MAJOR and MINOR classes of LP
Preparative Ultracentrifugation
108
(non-equilibrium or equilibrium
techniques) permits fractionation or several or ALL classes of LPs in a
single run
Density gradient methods -
109
Use antibody-coated plates specific for epitopes on apolipoproteins
both in routine and research lab
IMMUNOCHEMICAL METHODS
110
INDIRECT METHODS of LDL c estimation
friedewald equation
111
what is the equation of friedewald
LDL - TC - HDL - TG/2.175 in mmol/L
LDL - TC - HDL - TG/5 in mg/dl
112
Tedious -reserved for samples where Friedewald
equation is inappropriate
Beta-Quantification
113
(Reference method) for ldl estimation
Beta-Quantification
114
2 steps of Beta-Quantification
1. Ultracentrifugation to remove VLDL leaving behind LDL and HDL as well as IDL and Lp(a)
2. Chemical Precipitation of HDL-C
115
in the ultracentrifugation, we are removing ___ to keep __ and ___ as well
as IDL and Lp(a)
vldl; ldl and hdl
116
Limitations of ldl c estimation in indirect or friedwald method
not appropriate in:
1. Samples with TG > 400mg/dL
2. Patients with suspected Dysbetalipoproteinaemia
117
Other limitations of ldl estimation ; indirect or friedewald equation
1. Does not account for cholesterol associated with IDL and Lp(a)
2. Underestimate LDL-C in chronic alcoholics
3. Unsuitable for monitoring
4. Mis-classifies 15 - 40 % of patients when TG levels are between 200 to 400 mg/dL
118
Selectively measures LDL after masking non
LDL cholesterol, OR By selectively solubilizing LDL
Homogenous method
119
Advantages of homogenous method
Does not involve measurement of TGs thus
non-fasting samples can be used
Comparable to calculated LDL results from
beta-quantitation on normal lipemic
specimens
120
May be useful in evaluation of type Ill
hyperlipoproteinemia
VLDL-C/Plasma TG ratio
121
VLDL-C/Plasma TG ratio is expressed in
mol/mol or mass/mass
122
> 0.689 in VLDL-C/Plasma TG ratio means
hyperlipoproteinemi
123
VLDL-C/Plasma TG ratio ref range
0.689- 0.091
124
VLDL-C/Plasma TG ratio
Ranges ___ in samples without beta VLDL
0.230-0.575
125
give the arrangement of the lipoprotein from top to bottom arranged using electrophoresis
origin - chylomicron
ldl - beta
vldl - pre beta
hdl - alpha
126
the most anodic lipoprotein in electrophoresis
hdl - alpha lipoprotein
127
arrange the density of the lipoproteins out from ultracentrifugation
chylomicrons - 0.95 g/ml
vldl - 1.006 g/ml
IDL - 1.019 g/ml
LDL - 1.063 g/ml
HDL - 1.090 g/ml
fatty acids + albumin
128
in electrophoresis, we used the most sensitive type of gel or medium
agarose gel or polyacrylamide
129
stain used in electrophoresis for lipoprotein
Oil red O
Sudan
Fat Red 7b
130
former
Reference Method) for triglyceride
Van Handel and Zilversmith Method
131
in Triglycerides/TAG:
chemical method
1. Lipids are extracted using chloroform and
phospholipids and removed by __ absorption
zeolite
132
Principle or steps of van handel zilversmith method
1.
TAG -- > alcoholic KOH Glycerol + Fatty
acids
2. Glycerol + periodic acid --> Formaldehyde
3. Formaldehyde + Chromotropic acid ------> Blue
solution
133
in Van Handel and Zilversmith Method
what is added in the formaldehyde?
Chromotropic acid
134
in Van Handel and Zilversmith Method
what is the end product color
blue solution
135
a modification of in Van Handel and Zilversmith Method that will not use colorimetry but instead fluorimetry
hantzsch condensation method
136
hantzsch condensation method
steps or principle
1.TAG -- > alcoholic KOH -->Glycerol + Fatty
acids
2. Glycerol + periodic acid --> Formaldehyde
3. Formaldehyde + acetone + ammonia ------> yellow
solution
137
in hantzsch condensation method
what is added in formaldehyde
acetone and ammonia
138
in hantzsch condensation method
what is the end color
yellow
139
principle of enzymatic method of triglyceride is based on
glycerol kinase method
140
chromogen used in trinder's reaction in tryglyceridee determination
4 amino antipyrine
141
in chemical method - glycerol kinase method
the end color is
pink
142
Glycerol Kinase Method Read absorbance at ___ wavelength
500nm
143
Glycerol Kinase Method Read absorbance at 550 nm wavelength and linear up to
___
700mg/dL
144
INCREASED TAG are seen in
1. Hyperlipoproteinemia type i,
iib, iii, iv, v
2. Alcoholism
3. Nephrotic syndrome
4. Hypothyroidism
5. Pancreatitis
145
DECREASED TAG are seen in
1. MALABSORPTION
SYNDROME
2. Malnutrition
3. Brain infarction
4. Hyperthyroidism
146
an indicator of combined LDL and VLDL
concentration
Apo B:
147
major protein of HDL
Apo A-1:
148
the variant of LDL, an independent indicator
of CHD risk
Lp(a):
149
reference method or rnage of Apo A
120 - 160mg/dl
150
reference method or rnage of Apo B
< 120 mg/dl
151
reference method or range of Lp A
< 30 mg/dl
152
Hyperlipoproteinemias have been classified using the
system, which is not commonly used today.
Fredrickson-Levy classification
153
Mostcommonmethod for apoprotein
IMMUNOTURBIDIMETRY
154
easily adapted spectrophotometric analyzers allows the use of commercially available antisera and reference sera
IMMUNOTURBIDIMETRY
155
apoprotein that has indirect relation to coronary disease
apo A
156
other test we can conduct for apolipoprotein
ELISA , radial immunodiffusion, keme keme
157
Type I hyperlipoproteinemia: has an Elevated ___
chylomicrons
158
Type IIa hyperlipoproteinemia: has an increased
Increased LDL
159
Type lIb hyperlipoproteinemia: Increased ___
LDL and VLDL
160
Type III hyperlipoproteinemia: Increased ___
IDL
161
Type IV hyperlipoproteinemia: Increased ___
VLDL
162
Type V hyperlipoproteinemia: Increased ___
VLDL with increased chylomicrons
163
Total cholesterol level very low,
triglyceride level nearly undetectable, LDL and Apo B-100
absent
Abetalipoproteinemia
164
Unable to synthesize apo B-100
and apo B-48, low total cholesterol level and normal to low
triglyceride level
Hypobetalipoproteinemia
165
Severely elevated triglyceride
level and low HDL level
Hypoalphalipoproteinemia
166
HDL absent, apo A-I and apo A-II very low
levels, LDL low, total cholesterol level low, triglyceride level
normal to slightly increase
Tangier disease
167
