Immune sera are preparations derived from the blood of animals, usually horses, but mules, donkeys, sheep or goats are also used. The animals must be in good health, free of infections and obtained from sources free of transmissible spongiform encephalopathies, and kept under veterinary supervision.
IMMUNE SERA
A) Preparation
Immune sera
are preparations derived
from the blood
of animals, usually horses,
but mules, donkeys,
sheep or goats are also used.
The animals must
be in good health, free of infections and obtained from sources free of transmissible spongiform
encephalopathies, and kept under veterinary supervision. To prepare
an immune serum, horses or other animals
are injected with
a sequence of spaced doses of an antigen
until a trial
blood sample shows that
the injections have induced a high titre
of antibody to
the injected antigen. An adjuvant may be
used if required. A large
volume of blood
is then removed by venepuncture and collected into a vessel
containing sufficient citrate solution
to prevent clotting. The blood cells are allowed to settle and the supernatant plasma is drawn off.
Alternatively, the blood
can be mechanically defibrinated. The crude plasma can be sterilized by filtration and dispensed for use, but it is preferable to fractionate it to separate the immune globulin. This is done by
fractional precipitation of the plasma
by the addition
of ammonium sulphate. The globulin fraction is recovered and treated with pepsin to yield a refined immune
product containing the Fab fragment.
This refined globulin contains no more than a trace
of the albumin and other proteins that were present
in the plasma. It is less
antigenic, has a longer half-life in the circulation and is less likely to provoke
anaphylaxis or serum sickness than whole serum or crude globulin (Harms, 1948). The
antibody content
of the refined
product is determined by specific assay, the product is diluted to the required
concentration and transferred into ampoules. Two or more
monovalent immune sera may be blended together
to provide a multivalent immune
serum.
B) Quality Control
The quality
of immune sera
is controlled by potency tests and by conventional tests for
safety and sterility. The potency tests
have a common design in that, in the case of all immune sera,
the potency is estimated by comparing the
amount of the
product that is required to neutralize an effect
of a homologous toxin with
the amount of a standard preparation that is required
to achieve the same
effect. Serial dilutions of the immune
serum and of a
standard preparation are made and to each
is added a constant amount of the homologous antigen. Each mixture is then inoculated into a group of animals, usually guinea-pigs or mice,
and the dilutions of the immune serum and of the standard, which
neutralize the effects of toxin,
are noted. As the potencies of the standard preparations are expressed in IU, the potencies of the immune sera are determined in corresponding units
per millilitre (British
Pharmacopoeia, 2010).The quality of globulin fractions is usually monitored
by gel electrophoresis to detect
contaminating proteins and uncleaved
immunoglobulin,and by size-exclusion high-performance liquid chromatography to detect aggregates and small fragments. The immune
sera are also tested for contaminating viruses by inoculation on to cell cultures capable of detecting a wide range of viruses relevant
to the particular product.
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