Complement
o The complement system is a series of
more than 30 proteins normally found
in serum that play a major role in
phagocytosis and clearance of foreign
antigens from the body.
o The end product of complement
activation is lysis
o complement was so coined by Paul
Ehrlich
o Jules Bordet was awarded the Nobel
Prize in 1919 for his role in elucidating
the nature of complement.
o complement promotes
 opsonization and lysis of
foreign cells and immune
complexes,
 chronic activation can lead to
inflammation and tissue
damage
o Numerous proteins that act as controls
or regulators of the system.
 Most plasma complement
proteins are synthesized in the
liver
 C1 components, which are
mainly produced by intestinal
epithelial cells
 Factor D, which is made in
adipose tissue
 Other cells, such as monocytes
and macrophages, are
additional sources of early
complement components,
including C1, C2, C3, and C4.
o Most of the proteins of the complement
system are inactive enzyme precursors,
or zymogens, that are converted to
active enzymes in a very precise order.
 Activation of complement
o Classical pathway
 which involves nine proteins
that are triggered by antigen–
antibody combination
o Alternative pathway (properdin system)
 properdin’s major function is to
stabilize a key enzyme complex
formed along the pathway
o Lectin pathway
 antibody-independent means of
activating complement proteins
 major constituent, mannose- (or
mannan-) binding lectin (MBL),
adheres to mannose found
mainly in the cell walls or outer
coating of bacteria, viruses,
yeast, and protozoa.
 Complement system
 plays a major part in the
inflammatory response directed
against foreign antigen
THE CLASSICAL PATHWAY
o three main stages of Complement
activation
 The Recognition Unit
 C1 becomes activated when it
binds to the ends of antibodies
 C1q “recognizes” the fragment
crystallizable (FC) region of two
adjacent antibody molecules
 C1r cleaves a thioester bond on
C1s, which activates it
 C1s has a limited specificity,
with its only substrates being
C4 and C2
 Once C1s is activated, the
recognition stage ends
 o This pathway is important as an early
defense against pathogens
 Activation Unit o properdin does not initiate this pathway
 results in the production of an but rather stabilizes the C3 convertase
enzyme known as C5 convertase formed from activation of other factors
 C4 is the second-most- o C3, is a key component of both classical
abundant complement protein
and alternative pathways.
 C1s cleaves C4 to split off a 77-
o Triggering substances for the alternative
amino acid fragment called C4a
pathway
 C4b binds mainly to antigen in
 bacterial cell walls, especially
clusters that are within a 40 nm
those containing
radius of C1.
lipopolysaccharide;
 C2 is the next component to be
 fungal cell walls
activated
 Yeast
 The combination of C4b and
 Viruses
C2a is known as C3 convertase
 virally infected cells
 If C3b is bound within 40 nm of
 tumor cell lines
the C4b2a, then this creates a
 some parasites, especially
new enzyme known as C5
trypanosome
convertase
o C3bBb, one of the end products of this
 The cleaving of C5 with
pathway.
deposition of C5b at another site
o Factor D is a plasma protein that goes
on the cell membrane
through a conformational change when
constitutes the beginning of the
it binds to factor B.
membrane attack complex
(MAC)
LECTIN PATHWAY (mannose-binding, or
 Membrane Attack Complex mannanbinding, lectin (MBL)
 C5 consists of two polypeptide
chains, á and ß chain  binds to mannose or related sugars in a
 Subsequent binding involves calcium-dependent manner to initiate
C6, C7, C8, and C9 this pathway
o Membrane damage is caused by at least  produced in the liver and is normally
two different mechanisms present in the serum but increases
 channel formation and the during an initial inflammatory response
binding of phospholipids  plays an important role as a defense
o membrane attack unit mechanism in infancy, during the
 C6 binds to C5b, thereby interval between the loss of maternal
stabilizing antibody and the acquisition of a full-
 C7 binds next, forming a fledged antibody response to pathogens.
trimolecular complex that has a
high affinity for lipid
constituents of the cell
membrane
 C7 binds to C8 and binds to C9
 The completed MAC unit has a
functional pore size of 70 to
100Å.1,3 One such unit can
lyse erythrocytes, but lysis of
nucleated cells is a multihit
phenomenon.

THE ALTERNATIVE PATHWAY


Convergence of the classical, alternative, and
lectin pathways. The binding of C1qrs to two
antibody molecules activates the classical
pathway, while the alternative pathway is
started by hydrolysis of C3. The lectin
pathway is triggered by binding of MBP to
mannose on bacterial cell walls. MASP-1,
MASP-2, and MASP3 bind to form an
activated C1-like complex. MASP-2 cleaves
C2 and C4 and proceeds like the classical
pathway. Factor B and factor D operate in
the alternative pathway. While C3 convertase
is formed differently in each pathway, C3 is a
key component in each one. The C5
convertase in the alternative pathway
consists of C3bBb3bP. In the classical
pathway, C5 convertase is made up of
C4b2a3b. After C5 is cleaved, the pathway is
common to all.
Regulation of the Classical and Lectin
Pathways
o C1 inhibitor, or C1INH,
 is a glycoprotein with a
molecular weight of 105,000
that inhibits activation at the
first stages of both the classical
and lectin pathways.
 . Its main role is to inactivate C1
by binding to the active sites of
C1r and C1s.
 C1INH also inactivates MASP-2
binding to the MBL-MASP
complex,
 formation of C3 convertase in
the classical and lectin
pathways is inhibited by four
main regulators:
- soluble C4b-binding
protein (C4BP)
- complement receptor
type 1 (CR1)
- membrane cofactor
protein (MCP)
- decay accelerating
factor (DAF
o CR1, also known as CD35
 is a large polymorphic
glycoprotein with a molecular
weight between 165,000 and
280,000.
 It is found mainly on peripheral
blood cells
 2 It binds C3b and C4b but has
the greatest affinity for C3b.
 Once bound to CR1, both C4b
and C3b can then be degraded
by factor I Perhaps one of the
main functions of CR1
 immune adherence
 The ability of cells to bind
complementcoated particles
 The presence of DAF on host cells
protects them from bystander lysis and
is one of the main mechanisms used in
discrimination of self from nonself .
Regulation of the Alternative Pathway
o principal soluble regulator of the
alternative pathway is factor H
o Factor H also accelerates the
dissociation of the C3bBb complex on
cell surfaces
o When factor H binds to C3bBb, Bb
becomes displaced
• C3 convertase activity is
curtailed in plasma and on cell
surfaces.
o C3b in the fluid phase has a
hundredfold greater affinity for factor H
than for factor B
o factor H acts as a cofactor that allows
factor I to break down C3b
SYSTEM CONTROLS
Regulation of Terminal Components
o S protein also known as vitronectin is a
soluble control protein that acts at a
deeper level of complement activation.
o S protein interacts with the C5b67
complex
 o A receptor known by various terms,
including membrane inhibitor of reactive
lysis (MIRL), or CD59, also acts to block
formation of the membrane attack
complex
 its main function is to bind to
C8 and prevent insertion of C9
into host cell membranes
BIOLOGICAL MANIFESTATIONS OF
COMPLEMENT ACTIVATION
o anaphylatoxin
 is a small peptide that causes
increased vascular permeability,
contraction of smooth muscle,
and release of histamine from
basophils and mast cells
 Proteins that play such a part
are C3a, C4a, and C5a
 C5a- most potent; least 200
times more powerful than C3a;
serves as a chemotaxin.
 C4b,C5b,iC3b- opsonisation
COMPLEMENT DEFICIENCIES
Regulatory Factor Components
o paroxysmal nocturnal hemoglobinuria
(PNH)
 Individuals with this disease
have red blood cells that are
deficient in DAF
o DAF deficiency is associated with a lack
of CD59 (MIRL)
 CD59 has the same
glycophospholipid anchor found
in DAF
 CD59 prevents insertion of C9
into the cell membrane by
binding to the C5b678 complex
o Characterstic of Hereditary angioedema
 Recurrent attacks of
angioedema that affect the
extremities, the skin, the
gastrointestinal tract, and other
mucosal surfaces
LABORATORY DETECTION OF
COMPLEMENT ABNORMALITIES
Techniques to determine complement
abnormalities generally fall into two categories:
(1) measurement of components as antigens in
serum
(2) measurement of functional activity
Immunologic Assays of Individual
Components
o Radial Immunodiffusion (RID)
 uses agarose gel into which
specific antibody is
incorporated
o Nephelometry
 Measures concentration
according to the amount of
light scattered by a solution
containing a reagent
antibody and a measured
patient sample.
Assays for the Classical Pathway
 o The hemolytic titration (CH50) assay
 most commonly used for
this purpose
o lytic assays
 in general are complicated
to perform and lack
sensitivity
 Lytic activity can also be
measured by radial
hemolysis in agarose plates
o ELISAs
 another means of measuring
activation of the classical
pathway
 C4a, C4d, C3a, and C5a, which are
generated only if complement activation
has occurred

Interpretation of Laboratory Finding

Complement Fixation Testing

o Complement itself can actually be used

as a reagent in the test known as
complement fixation