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The Biologic Structure and Function of CRP |
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Tillet and Francis first discovered CRP in 1930 at Rockefeller University while examining the serum of adult
patients diagnosed with acute pneumococcal pneumonia. They observed a precipitation reaction between CRP and the
C-polysaccharide cell wall of the pneumococcal bacteria. This reaction later was determined to be the result of
CRP binding to C-polysaccharide in the presence of calcium, forming CRP-ligand complexes. CRP's unique binding
characteristics have led to the identification of elevated CRP levels in over 70 different infectious and noninfectious
disorders associated with acute and chronic inflammatory disorders in adults, children, and infants.
Understanding the biochemical structure and function of CRP is important to ensure appropriate use. CRP is a member
of the pentraxin family of proteins, which are nonspecific, acute-phase reactant proteins composed of 5 identical
23-kDa polypeptide subunits arranged in a cyclic pentameter shape.] Each of these subunits contains one binding site
for a phosphocholine molecule and 2 binding sites for calcium.
CRP as an Acute-Phase Protein
An acute-phase inflammatory response occurs as the human body mounts a systemic response to tissue injury caused by
infectious, noninfectious, chemical, physical, or immunologic toxins. CRP, serum amyloid A, haptoglobin, and fibrinogen
are all positive acute-phase proteins regulated by cytokines, which are known to increase by at least 25% during the
inflammatory response.[32] Interleukin-1 (IL-1), interleukin-6 (IL-6), and tissue necrosis factor (TNF)- are the major
cytokines that stimulate the liver to synthesize CRP and other positive acute-phase proteins
CRP is the most sensitive of the acute-phase proteins, with levels rising as much as 1,000-fold during the acute
inflammatory processes. Levels begin to rise within 4 to 6 hours of the onset of signs of infection or tissue injury
and peak 24 to 48 hours later. They rapidly disappear as the infection or inflammatory process resolves.
The degree of CRP response or rise in serum may be dependent on the amount of tissue damage present. For example,
elevated CRP levels from 150 to 350 mg/L have been reported in cases of invasive bacterial meningitis, whereas smaller
rises from 20 to 40 mg/L occur in acute viral infections and from noninfectious causes. CRP is a useful serum marker to
assess and monitor the presence, severity, and course of the inflammatory response in infectious and noninfectious
disorders, including acute myocardial infarction, angina, malignancies, rheumatoid arthritis, inflammatory bowel disease,
burns, and trauma, and after surgical procedures.
Laboratory Methods to Measure CRP
The 3 clinical laboratory methods used to measure serum CRP levels are as follows:
- Qualitative
- Semiquantitative
- Quantitative
All 3 tests are based on the ability of CRP to bind to a variety of biologic ligands forming CRP-ligand complexes.
When a reagent containing anti-human CRP antibodies is added to a serum sample containing CRP, the CRP binds to the
antibodies forming insoluble CRP-ligand complexes that clump and precipitate, which can then be visualized and measured.
Qualitative Testing Method
The qualitative latex agglutination test is the first laboratory method developed to measure CRP. This method measures
the presence or absence of agglutination and precipitation, indicating only whether CRP is present or absent in the
serum sample. A positive test result indicates the presence of CRP-ligand complexes formed when CRP binds to cause
agglutination and precipitation, whereas a negative test result occurs when no agglutination is present. Positive test
results indicate a CRP level greater than 6 mg/L or more than 10 mg/L, depending on the specific testing kit and reagent
being used Qualitative tests can be performed rapidly at the bedside within 10 to 15 minutes.
Purely qualitative CRP testing methods typically are not used to measure CRP levels because they have a low sensitivity,
with positive results occurring with any condition involving inflammation or tissue destruction.[12, 29, 35] A positive
qualitative CRP test should always be followed by a semiquantitative test, which is a more sensitive measuring method to
quantify the concentration of CRP.
Semiquantitative Testing Method
The semiquantitative latex agglutination assay involves the use of serial dilutions of serum and saline. Each is mixed
with a latex reagent and observed for the presence of agglutination.[29, 44, 45] The highest dilution in which
agglutination is visualized corresponds to an approximate titer or concentration of CRP-ligand complexes.
Like the qualitative test, the semiquantitative test can be performed in 15 to 30 minutes and has a reported upper
detection level between 6 and 10 mg/L.[35, 45] Results are reported as a ratio and/or an approximate mg/L concentration
of CRP.[44, 45] The reported cost of a Murex semiquantitative testing kit (Murex Diagnostic, Norcross, GA) is
approximately $21.00.
Quantitative Testing Method
Quantitative immunoassay is the most rapid, sophisticated, and sensitive method of detecting and measuring CRP.[]
The enzyme-linked immunoabsorbent assay (ELISA) and immunofluorescence quantitative test uses monoclonal anti-CRP
antibodies marked with an enzyme or fluorescent tracer fixed to microwells located within a microtiter plate or test
tube. When diluted human serum is added, CRP binds to the immobilized marked anti-CRP antibody forming bound CRP-ligand
complexes. Unbound antibodies are then washed from the test tube. The fluorescent-marked CRP-ligand complexes can be
visualized and measured under a fluorescence microscope. A substrate is added to the enzyme-marked CRP-ligand complexes,
which reacts with the enzyme causing a color reaction. The specimen is read with a spectrophotometer, which calculates
the net color absorbency. This method of testing can be performed within 10 to 15 minutes and is capable of detecting
CRP levels between 1 to 40 mg/L.
Laser and rate nephelometry quantitative immunoassays use infrared light-emitting diodes and detectors passed through
test tubes containing fixed amounts of anti-CRP monoclonal antibody mixed with human serum, resulting in the formation
of CRP-ligand complexes. Because the antibody concentration remains constant, the extent of light scatter is determined
from the amount of CRP-ligand complexes. This method can be performed within 30 to 60 minutes. When using a fully
automated analyzer, however, it takes 10 minutes. The cost of the fully automated quantitative CRP test is reported to
be around $50.00.
Qualitative and semiquantitative methods of measuring CRP, although less expensive, are not accurate enough to be used
for infants.[12, 27, 29] The limited detection level of 6 mg/L in some kits could easily miss infants with true sepsis
and result in longer antibiotic therapy.[47] It is imperative that practitioners be aware of which CRP testing method
is used in their own institution and the sensitivity and specificity of that test when evaluating measured CRP levels
in the clinical setting.
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