C-REACTIVE PROTEIN AS A

DIAGNOSTIC TOOLS FOR

ASSESSING ATHEROSCLEROSIS
BY

SHUAIBU BUKHARI

3RD NOVEMBER , 2017

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 OUTLINES

 INTRODUCTION

 STRUCTURE /FUNCTIONS OF CRP

 CRP & ATHEROSCLEROSIS

 CRP AS A SCREENING TOOLS

 METHODS/PRINCIPLES OF ASSAYING CRP

 INTERPRETATION OF RESULTS

 CONCLUSION/RECOMMENDATION

 REFERENCES
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 INTRODUCTION
 C-reactive (CRP) also known as pentraxin is a 224 amino acid protein that is
synthesized primarily by hepatocyte and released into the blood within a few
hours after tissue injury.

 CRP was first described in 1930 by Tillet & Francis, named after its ability to
precipitate & interact with phosphocholine residues of the C-polysaccharides
derived from techoic acid within the cellullar wall of strptococcus
pneumoniae, as well as its ability to precipitate with calcium (Kind & Pepys,
1984).
 CRP is one of the substances present in the atherosclerotic lesion, more
specifically in the vascular intima where it co-localizes with monocytes &
Lipoproteins
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Atherosclerosis also known as arteriosclerotic vascular disease(ASVD) is

a process by which the artery wall thickens as a result of invasion &

accumulation of WBCs & proliferation of internal-smooth muscle cell

creating an atheromatous (fibrofatty plaque)

The fibrofatty plaque is made up of fats, CRP, cholesterol, calcium &

other substances found in the blood

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 STRUCTURE / FUNCTION OF CRP

Recognition of ligands

Activation of complement via classical

pathway
Ca2+ are necessary for
lagand binding Binding to receptors on phagocytic
Phosphocholine in the
cells/enhancement of phagocytosis
ligand binding site
Induction of cytokines synthesis
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 CRP & ATHEROSCLEROSIS
 CRP has been implicated as a contributor to atherogenesis by modulating
endothelial function, stimulating coagulation, inducing expression of 1CAM-1,
VCAM-1, & mediating uptake of LDL into macrophages (Deferranti & Rifail,
2002).

 CRP binds to modified forms of LDL, & when aggregated, CRP can bind to
native LDL as well.

 Most clinical studies report that CRP is an independent predictor of risk of:
atherosclerosis, cardiovascular events, atherothrombosis, hypertension &
myocardial infarction 1/8/2020
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Framingham, (2002) suggest that the C-reactive protein level is a
stronger predictor of cardiovascular events than the LDL cholesterol
level.

Another study suggest that CRP levels may be useful in identifying

apparently healthy men who are at an increased long-term risk of
sudden cardiac death (Circulation ,2002)

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 CRP AS A SCREENING TOOL
C-reactive protein (CRP) is an acute phase marker and a predictor of the
risk of developing atherosclerotic complications.

High sensitivity CRP has been used as a means of assessing

atherosclerosis.

Therefore, it has been suggested that risk assessment should not rely
purely on LDL-C measurements because there is evidence that CRP had
a stronger predictive value for the risk of CHD events than LDL-
C(Ridker, 2008).
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 Method of Assaying CRP

High-sensitive Nephelometric method

Turbidity metric method

immunometric Assay

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 Principle of Nephelometric method & Turbidity Metric Method
 The CRP reacts with the specific antibody producing insoluble immune
complexes. The turbidity caused by these immune complexes is proportional
to the CRP concentration in sample and can be measured
spectrophotometrically.

Procedure
 Mix Serum (or heparinized plasma) with Intralipid 20% in Tris-calcium buffer
(pH 7.5).
 Incubate for 12min(Nephelometric method) & 30min (Turbidity metric
method) at 37°C respectively
 Measure the CRP-phospholipid complexes by nephelometry (840 nm) using a
BN II nephelometer (Siemens) or turbidimetry (660 nm/700 nm) with a
Cobas
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 PRINCIPLE OF ELISA ASSAY FOR CRP
 This immunometric Assay is based on double –antibody
“Sandwich technique.

 Each wall of the micro well plate supplied with the kit has
been coated with a mono-clonal antibody specific For Human
CRP (Mouse Anti-Human CRP)

 This antibody will bind any CRP introduced into the well.

 Standards and samples are incubated on the antibody –coated

plate and the plate is then Rinsed before addition of a HRP
labeled CRP monoclonal antibody to detect the captured
CRP

 The two antibodies formed a Sandwich by binding to

different location on the CRP molecules

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 The concentration of the analytes is determined
by measuring the enzymatic activities of HRP
Using a chromogenic substrate TMB(3,3,5,5
Tetra Methyl Benzidine)

 After a sufficient period of time, the reaction is

stopped with acid forming a product with
distinct Yellow Color that can be measure at
450nm

 The intensity of this color, determined

Spectrophotometrically, is directly proportional
to amount of bound HRP labeled monoclonal
antibody which in turn is proportional to the
concentration of the CRP

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 INTERPRETATION OF RESULTS
A C-reactive protein is measured in milligrams of CRP per liter of blood
(mg/L).

According to CDC, a reading of less than 1 mg/L indicates low risk of

cardiovascular disease.

A reading between 1 and 2.9 mg/L means an intermediate risk.

A reading greater than 3 mg/L means high risk for cardiovascular

disease.

A reading above 10 mg/L may signal a need for further testing to

determine the cause of such significant inflammation in your body.
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 Conclusion/Recommendation

Elevated levels of CRP may have an important role in identifying those

who may be at risk of atherosclerosis.

CRP should be analyzed along side with LDL-cholesterol to uncovered

those at risk for heart disease where cholesterol levels alone may not be

helpful.

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 REFERENCES

Pearson TA, Mensah GA, Alexander RW, et al. Markers of Inflammation and Cardiovascular
Disease: application to clinical and public heath practices: A statement for healthcare
professionals from the Centers of Disease Control and Prevention and the American
Heart Association. Circulation 2003: 107; 499-511

Framingham,N .(2002)Engl J Med 14;347(20):1557-65

Circulation (2002 ). C-reactive protein 4;105(22):2595-9

Factfile BHF.(2007). Novel Risk Factors and the prediction of Coronary Heart Diseas Risk

Kind CRH, Pepys MB. The role of serum C-reactive protein(CRP) measurement in clinical
practice. Int Med 1984;5:112-151.

Gerwurz H, Mold C, Siegel J, Fiedel B. C-reactive protein and the acute phase response. Adv
Intern Med 1982;27:345-71.
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