3. Biochemical Tests Are
Required For
a-
Screening
b- Diagnosing
c- Monitoring or Management
d- To establish prognosis
3
4. ACTUAL ANALYTICAL
PROCEDURE
A-
PRE-INSTRUMENTAL PHASE
1- Selection of Patients
2- Prepration of Patients
3-Collection of Sample
4- Transportation of Sample
B- INSTRUMENTAL PHASE
1-Methodology
2-Calculation of Values.
C- POST INSTRUMENTAL PHASE
1-Reporting
2-Evaluating the reports with Patients
clinical stage.
4
6. INTERNATIONAL VALUES
1-
International Union of Pure & applied
chemistry.(IUPAC)
2- International Federation of clinical
chemistry.(IFCC)
3- Systemic International units.(SI)
These units help us to communicate the
results to larger scientific community with
clearity.
6
8. EXERCISE
A
TRANSIENT EFFECT-(within an hr.)
-These are due to increased metabolic activity
eg.-Free Fatty Acids-
-Alanine conc.-180%
-Lactate
-300%
-They are corrected to pre-exercise level soon
after the cessation of the exercise.
8
10. PREVIOUS DIET
Usually
biochemical analysis is recommended in
fasting stage.(12 hrs.)
Prolonged Fasting- 24hrs. Fasting can lead to an
increase in - Serum Bilirubin
- Fatty acid
- Valine, Leucine
> 72 hrs. fast can lead to decrease in
- Plasma –glucose
-protein,albumin
-pre-alb,transferrin
10
11. After Meal- Physiological effects of eating a meal
within 2-4 hrs. includes increase in
-Triglycerides
-Potassium
In vivo changes are dependent upon the type and
quantity of food ingested eg. 1-High fatty diet:- increases
-Intestinal isoenzyme of alkaline phos.sp.
In ‘O’-B’ group
-Turbidity- LACTESCENT SERUMWhich can interfere in many assays.
2-High Protein diet;- can cause increase in
-Blood Urea,Ammonia, Urate etc.
3-High Ratio Unsaturated: Saturated Fats causes
decrease in CHOLESTEROL.
11
12. Diet
rich in Purine:- Increases
- Urate conc
Banana,Pineapple,Tomato. Because of presense of
serotonin can cause increase in
Renal 5 Hyd roxyindol acetic acid .
(Diagnostic-Carcinoid)
Coffee:- causes threefold rise in
-Nonesterified free fatty acids.
-Cholestrol
-Cause a release of catecholamine
from the adrenal medulla.
Ethenol;-1.-Immediate changes are increase in
Lactate,Urate, Acetaldehyde,Acetate
2.-Intermediate changes ( 10-100HRS.)
(mainly on serum enzymes)
12
14. Long
time effect:- Increase in Triglyceride
It has been noted that chronic alcoholics
have higher plasma HDL cholesterol conc.than do
matched control subjects. Abusers of alcohol show
increased value of serum GAMMA GLUTAMYL
TRANSFERASE,URATE &MEAN
ERYTHROCYTE VOL.
Tobacco smoking:-results in increase in the % of
blood Carboxy Hb
Acute effects of Tobacco smoking include
increase in plasma catecholamine and serum
cortisol.(Probably due to the nicotine)
Chronic effects are increase in Hb,MCV,TLC
14
15. EFFECT OF DRUGS
Young
had studied about 15,000 drugs and
found that they have
A.- Physiological effect.- In vivo effects of
the drug or its metabolites.
B.- Chemical interference.- In vitro due to
some physical or chemical property which
interfere with the assay.
15
19. A
no. 0f drugs are known to affect the liver by
inducing hepatic microsomal enzyme or by
producing hepatocellular damage or cholestatic
jaundice.eg.
-Synthetic steroid
-Sulfonylurea derivatives eg.
Chlorpromazine
Thiouracil
Tolbutamide
- Erythromycin
-INH
- Tetracycline
-Aspirin
Cholestatic changes are increase in Alk.Phos.,Bromosulphalein,Bilirubin,AST/ALT
19
20. Oral
contraceptives:-Physiological effects include
increase in-ALT
- Ceruloplasmin
- Transcortin
-Thyroxin binding globulin
- Iron
- Triglyceride
-Gamma Glutamyl Transferase
decrease in
-Albumin
-Zinc
These effects are probably related to the estrogen
portion of the drug.
20
21. POSTURE
Differ
OPD:INDOOR
Samples are usually taken in either supine or
sitting position.As patient goes from supine to
standing posture there will be an efflux of water
and filterable subs.from I/V to interstitial fluid
space.Non-filterable subs.like protein and protein
bound subs.are increased in blood.There is sig.
Change in supine to standing stages. STATLAND:- Within 30 mints.there is an
increase in:21
23. EFFECT OF TOURNIQUET
OR FIST EXERCISE
Prolonged
application of tourniquet or fist
exercise cause significant changes in the conc.of
many serum constituents eg.
-Lactate
-Enzymes
-Protein
-Protein bound subs.such as
Cholestrol
Triglyceride
Calcium&Iron
23
24. Statland
compaired 1mt.:3mt.Tourniquet
application and found an increase in
Protein
5%
Cholesterol 5%
Iron
6%
Bilirubin
8%
AST
10%
STRESS:Effects Adrenal Hormones.i.e.why Cortisol
estimation is done at 8AM&8PM
ANXIETY Hyperventilation leads to
disturbance in acid base balance.An increase in
- Lactate
-Non-esterified fatty acid
24
25. ERRORS DUE TO
PREPRATION OF SPECIMEN
-During collection
-Specimen interferance
-During processing of specimen
-During storage
25
26. COLLECTION OF
SPECIMEN
A.Identification error:-a. Proper Reqesition Form
b.Name,CR No.,Bed No.etc.
c.Collection time in 24hrs.
specimen.
d.Correct type of container
-Anticoagulant
-Preservative
B.Site of Blood drawing:a.Postural
b.Tourniquete
c.Capillary or venous
26
27. Technician
should be certain to avoid sampling
from an extremity in which an I/V catheter is
delivering parentral soln.
Site of venipuncture must be distal to the I/V
needle and the tourniquete must be placed
between the I/V needle and the site of
venipuncture.
Choice of capillary or venous blood is not usually
of clinical importance except in the case of GTT
where capillary conc. is 10-30% higher
C.Contamination of specimen:1.Residual detergent:-May contaminate
the sample with Inorganic phosphate
2.Plasticizers:-(I/Vset,Tube,Rubber
cork)create spurious peaks in gas liquid
chromatography.
27
28. 3.Glass tube,Cork etc.-Inceases
calcium.
4.Container with lead.-Increases Lead
5.Metal of needle:-Interferes with
-Assay of coagulation
-Platelets.
Haemolysis;-While venipuncture
-When blood collected in vaccum tubes
-While mixing
-Greater bore needle has more chances
28
29. The
majority of chemical measuremants are
performed on specimen obtained from the
e.c.f.ie.usually plasma or serum.Certain analytes
are present in the formed elements of the blood in
conc. many times higher or lower than in the
surrounding plasma and therefore lysis of the cells
will contaminatethe plasma or serum to a
measureable amount.
Direct interference of Hb.increases:-Bilirubin
-Protein
-Potassium etc.
Haemolysis can occur before venipuncture or
during the analytical procedure.
29
30. 1% of Haemolysis can change:LDH
+272%
AST
+220%
POTASSIUM +25%
ALT
+55%
GLUCOSE
--5%
INORGANIC PHOS. +9.1%
SODIUM
--1%
CALCIUM +2.9%
ICTRIC SERUM:-Jaundice of 2.5mg./dl will
interfere with estimation of
-Cholesterol -Glucose -Albumin
Can be eliminated by using appropriate blanking
procedure or dual wavelength.
30
32. EFFECT OF
ANTICOAGULANTS
A.-Potassium
oxalate or EDTA:-Causes decrease
in
-Calcium
-Amylase
-LDH
B.-Floride:- Inhibits:-Glucose oxidase
-Acid phosphatase
-Amylase
So preferrably serum is used for biochemical tests
32
33. A.
CHANGES DURING
STORAGE
Evaporation:-Of water from serum
-Results in higher conc.of all
analytes
-Causes increase in activity of
enzymes
10mts.storage will cause 5% increase in osmolality at
28 degree C Temp.with 25% Humidity.
B.Refrigeration:-Of amniotic fluid increases
- Lecithin: Sphingomylin
-Phospholipase decreases
33
34. C.-Open
Sample:-I.Will lead to evaporation of
CO2 which increases pH to 8.5 in 2hrs.which
causes decrease in Acid phosphatase.
2.Glycolysis- Decreases Glucose.
3.Proteolytic&Hydrolytic processes increase conc.
Of Ammonia.
4.Changes in Erythrocyte Permeability increases
- Potassium
-Phosphate & Magnesium
D.-Exposure to light:-Decreases
-Bilirubin
-Delta aminolevulinic acid
-Porphyrins
-Porphobilinogen.
34
36. Idealy
all measurements should be performed
within 1hr.after collection. Prolonged contact of
serum with cell clot beyond two hrs.can cause sig.
Changes in certain constituents.Such as
Glucose,Potassium,Phosphorus,Creatinine,AST
&ALT(Rehak).
Clinically useful data generated by the lab must be
reported promptly and accurately to optimize
patient management.Delay in reporting can make
data useless.
36
37. QUALITY CONTROL
Quality
control is must in the lab.
-Interlaboratory
-Intralaboratory
Interlaboratory:-Standards
-Referance samples-Low
-High
37
38. EVALUATING RESULTS
Before
evaluating results we should check
-Reliability of Method
-Specificity&Senstivity
-Random Analytical Variation
-Dynamic Range
-Interferance
38
39. CONCLUSION
Total
variation in results of patients can be
grouped:
- Analytical Variation
-Prepration of subjects
-Intra Individual Physiological
-Inter Individual Biological
variations of Mean Values.
39
40. One
has to appreciate and keep in mind
these expected non pathological source of
variation in Diagnosing and management of
Disease.
40
