IMMUNOSERO LAB
microorganism
C-REACTIVE PROTEIN DETERMINATION
 Acute phase reactant present in normal
serum (fibrinogen,complement proteins
<haptoglobin>)
 during inflammation, they change
concentration (increases)
 to check the level of crp if detectable or
not
 Liver: main organ necessary for the
production of most proteins
 Pentameric acute phase (5
molecules/monomer attached to each
other)
 Acute phase of pneumococco infection
in 1930
 Derived from c-polysaccharide of
pneumococci
 Innate immunity
METHODS
- Routine test (qualitative test)
- Pentraxin group of the plasma proteins
(five identical peptide subunits)
- Half-life of crp: 18 hours increases
rapidly within 4-6 hours after infection
- Reaches peak value within 48 hours
(hundred or even thousand fold)
- Widely used indicator of acute
inflammation
- To observe response of treatment
- Can also use esr in respond to
inflammation
- Provides direct serum protein
measurement
- Crp is easily destroyed by heating the
serum 56C for 30 minutes
- Plasma half-life:
- Main substrate: phosphocholine
(presence of calcium, crp binds to
polysaccharide) calcium dependent
- Activates classical complement pathway
- Phosphocholine found surface of
- Necessary because it interfere with
some testing like detection of
antibodies
- Production of crp is in the liver under
the control/stimulation of Interleukin-
6(pyrogen) only il that activates apr
(important mediator of fever and acute
phase response)
- il are cytokines (signaling proteins of
phagocytic cells) subclass are
interleukin
- B cells, hepatocytes,
- Excessive production of il-6 is
dangerous (excessive production:
cytokine storm) during chronic
inflammation
- Crp is also an opsonin that activates
complement classical pathway that
leads to lysis.
- Normal level in male: appx. Less than
1.5mg/L
- Normal level in female: appx. less than
2.5mg/L
Pentameric crp (phagocytosis) dissociate in the
site of inflammation and becomes monomeric
crp which promotes chemotaxis
Crp stimulates immune cells through binding to
the fc receptor of the antibody which increase
the production of immunoglobulin
70 diseasse state
Extremely elevated level crp – possible bacterial
infection in at risk patients
Crp is not specific to rule out the cause of
increased in crps
Prior to chemoteraphy,, if crp is higher than
10mg/L, if decrease by 50% after 2 seconds,
chemoteraphy is effective and found related to
cancer.
When there is a rejection in organ/tissue
rejction donated, rcp levels will rise
After surgery, it will peak after 2 days and
gradually return after 7-10 days but if crp is
persistently elevated, patient has underlying
sepsis
For obese patient, crp and ldl cholesterol is
present (ldl transports cholesterol from the liver
to the tissue) hdl should always be more than
ldl
People at risk in cardiovascular diseases
Myocardial infarction (atherosclerosis/clog of
blood) stroke (brain/cerebrovascular accident)
Cdc recommended the crp concentration of less
than 1 mg/L is associated with low risk of
cardiovascular disease
1-3mg/L average risk
3 mg/L and above high risk for hjeart attack and
stroke
More than 10 mg/L Abnormally high (if you
have severe infection)
CRP rapid latex agglutination test
- Carrier: latex particles coated with
antihuman crp antibodies against
unknown serum which binds to human
crp in the serum
- Agglutination, above the reference
interval
- More than 6 mg/L if reactive (analytical
sensitivity)
- Principle: reverse passive agglutination
(detecting antigens), slide test or
qualitative test
- Reagent: 1 percent suspension of
polystyrene latex particles coated with
anti-human crp produced in goats or
rabbits
- Saksak human crp sa rabbit, will
recognize as foreign so magpproduce ng
produce anti-human crp
- If passive, carrier is attached to antigens
(detect antibody)
- Preservative: sodium azide (highly toxic
and corrosive)
- Specimen: fresh and clear serum
- anti human crp antibody IGG
- Glycine saline buffer 20%/ 1 glycine
saline : 20 water (8.1 to 8.3 slightly
alkaline solution)
- Sodium azide: 0.1% (0.96g/L)
- Mix the vial, gitna sa palm
- Quality control to know if agglutination
is produced, positive.
- Positive control: human serum with crp
concentration of .15 mg/L & 0.95mg/L
sodium azide
- Negative control: animal serum with
max. concentration of human crp of
1mg/L & 0.95mg/L sodium azide
- Undilutes patient serum
- 1 drop of patient serum, hold pipette
perpendicularly to the slide because the
amount of serum being dispensed will
not be the same (at least 50 microliter)
then add 1 drop of positive control to
the positive circle and 1 drop of
negative control to the negative circle
then suspend the latex reagent. Mix the
vial carefully. Lagay ng latex reagent sa
tabi lang ng bawat drop, then mix with
separate stirrer for each mixture and
maximize until it reaches/fill up the
mark of the cicle. Tilt test slide back and
forth or 100 rpm if mechanical rotator
then observe for agglutination OF
LATEX SUSPENSION (mostly on the ring
part)
- Positive reaction: agglutination of Latex
Suspension (if crp concentration is
higher than 6mg/L)
- Negative Reaction: opaque / smooth
milky suspension
- Antigen excess: postzone so dilute the
serum sample
-

- Centrigufe to remove upper lipids if

lipemic.
- False negative: aspirin (anti-
inflammatory)

SEMI QUANTITATIVE METHOD

- Dilute the serum/specimen; titrate the

amount of crp (6 test tubes)
- Most diluted is tube number