Biomedical Engineering Reference
In-Depth Information
Antibody preparations used to induce passive immunity may be obtained from either animal or human
sources. Preparations of animal origin are generally termed 'antisera', and those sourced from humans
are called 'immunoglobulin preparations'. In both cases, the predominant antibody type present is IgG.
Antisera are generally produced by immunizing healthy animals (e.g. horses) with appropriate
antigen. Small samples of blood are subsequently withdrawn from the animal on a regular basis
and quantitatively analysed for the presence of the desired antibodies (often using ELISA-based
immunoassays). This facilitates harvesting of the blood at the most appropriate time points. Large
animals, such as horses, can withstand withdrawal of 1 or 2 l of blood every 10-14 days, and anti-
body levels are usually maintained by administration of repeat antigen booster injections.
The blood is collected using an aseptic technique into sterile containers. It can then be allowed
to clot with subsequent recovery of the antibody-containing antisera by centrifugation. Alterna-
tively, the blood may be collected in the presence of heparin, or another suitable anticoagulant,
with subsequent removal of the suspended cellular elements, again by centrifugation. In this case,
the resultant antibody-containing solution is termed 'plasma'.
The antibody fraction is then purifi ed from the serum (or plasma). Traditionally, this entailed
precipitation steps, usually using ethanol and/or ammonium sulfate as precipitants. The precipi-
tated antibody preparations, however, are only partially purifi ed and modern preparations are
generally subjected to additional high-resolution chromatographic fractionation (Figure 13.1).
Ion-exchange chromatography is often employed, as is protein A affi nity chromatography. (IgG
from many species binds fairly selectively to protein A.)
Following high resolution purifi cation, the antibody titre is determined, usually using an appro-
priate bioassay, or an immunoassay. Stabilizing agents, such as NaCl (0.9 per cent w/v) or glycine
(2-3 per cent w/v) are often added, as are antimicrobial preservatives. Addition of preservative is
particularly important if the product is subsequently fi lled into multi-dose containers. Phenol, at
concentrations less than 0.25 per cent, is often used. After adjustment of the potency to fall within
specifi cation, the product is sterile fi ltered and aseptically fi lled into sterile containers. These are
sealed immediately if the product is to be marketed in liquid form. Such antibody solutions are
often fi lled under an oxygen-free nitrogen atmosphere in order to prevent oxidative degradation
during subsequent storage. Such a product, if stored between 2 and 8
C, should exhibit a shelf life
of up to 5 years.
Although specifi c antisera have proven invaluable in the treatment of a variety of medical con-
ditions (Table 13.1), they can also induce unwanted side effects. Particularly noteworthy is their
ability to induce hypersensitivity reactions; some such sensitivity reactions (e.g. 'serum sickness')
are often not acute, whereas others (e.g. anaphylaxis) can be life threatening. Because of such
risks, antibody preparations derived from human donors (i.e. immunoglobulins) are usually pre-
ferred as passive immunizing agents.
Immunoglobulins are purifi ed from the serum (or plasma) of human donors by methods similar
to those used to purify animal-derived antibodies. In most instances, the immunoglobulin prepara-
tions are enriched in antibodies capable of binding to a specifi c antigen (usually an infectious mi-
croorganism/virus). These may be purifi ed from donated blood of individuals who have recently:
•
been immunized against the antigen of interest;
•
recovered from an infection caused by the antigen of interest.
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