The term antibody fragments covers a large number of molecular structures derived from full-length antibodies. Some clarification
of structural types is useful before discussing production aspects.
IgGs are the most abundant immunoglobulins in the blood and are commonly used as antibody therapeutics. They have a molecular
weight of approximately 160 kDa comprising 12 domains of roughly equal size. They are made up of four separate peptide chains,
two identical heavy (H) chain polypeptides and two identical light (L) chain polypeptides (Figure 1). The L and H chains are
composed of two and four domains, respectively, with each domain having a similar β-barrel structure and containing one disulfide
bridge. The H and L chains are linked together by a combination of disulfide and non-covalent bonds.4 Each chain contains both variable and constant domains. The variable domains of the H and L chain (VH and VL) contain the
variable complementarity-determining regions (CDRs). These regions of extremely variable amino acid sequences are located
at the N-terminal part of the antibody molecule. Together, VH and VL form the unique antigen-recognition site. The amino acid
sequences of the remaining C-terminal domains are much less variable.
Figure 1. An explanatory schematic of whole antibody domain structure
The Fc region, also referred to as the constant domain, is the nonantigen-binding part of the antibody molecule. Fc mediates several immunological functions, such as binding to receptors
on target cells and triggering effector functions that eliminate the antigen. There are applications where the Fc-mediated
effects are not required or are even desirable. The Fc fragment also is the site of glycosylation in IgGs.
The unique antigen-binding site of an antibody consists of the H and L chain variable domains (VH and VL). Each domain contains
four conserved framework regions and three CDR regions, which determine the specificity of the antibody. The VL and VH domains
together form a binding site, which binds a specific antigen.
A wide variety of functional antigen-binding fragments has been constructed from whole antibody domains, some of which are
illustrated in Figure 2 (see also reference 5). Selected fragment types are discussed in detail below.
Figure 2. Antibody fragment diversity
Fab fragments (fragment antigen binding) are the antigen-binding domains of an antibody molecule, containing two amino acid
chains composed of two domains, VH + CH1 and CL + VL. An interchain disulfide bond present at the C terminal of each constant
domain links the CL and CH1 domains together, which gives a molecular weight of the heterodimer around 50 kDa.6 Lucentis and Cimzia are both examples of Fab antibody fragments.
Single-Chain Fv Fragments
A single-chain Fv fragment (scFv) is the smallest fragment (~30 kDa) that still contains the complete antigen-binding site
(VH + VL) of a whole IgG antibody.7 Linker sequences joining the VH and VL domains in a single amino acid chain have been used to overcome the fact that native
scFv fragments are unstable and tend to dissociate from one another.
Antibody Fab and scFv fragments, comprising both VH and VL domains, retain the specific monovalent antigen-binding affinity
of the parent IgG while showing improved pharmokinetics for tissue penetration. However, these are not the smallest form of