VH or Vκ genes amplified from each single cell are cloned into IgG1 or Igκ

expression

vectors.

Heavy-

and

light-chain

plasmids

are

subsequently

co-transfected into the mammalian cell line such as HEK 293A cells for expression

of the antibody. Expression of specific human antibody in the culture supernatant is

confirmed by using high-throughput screening assays. Subsequently, antibody can

be purified by using protein A/G Sepharose column. A large number of human

antibodies against different neutralization-relevant epitopes of the glycoprotein of

Ebola virus have been isolated from the survivor of the 2014 Ebola outbreak

(Bornholdt et al. 2016). Similarly from influenza vaccinated human subjects, using

single-B cell PCR, approximately 50 human MAbs binding to 3 influenza vaccine

strains with high affinity have been generated (Wrammert et al. 2008).

22.5

Various Formats of Therapeutic Antibodies

Depending on the clinical application of the therapeutic antibodies, beside full-

length antibodies, these have been expressed in various formats (Fig. 22.4), which

are briefly described below:

VH

VL

scFv

Antibody

VH

VH

VL

VL

Bi-specific

Antibody

Pro-drug

Drug

Antibody-drug

Conjugate

VH

VL

CH

CL

Fab fragment

Fig. 22.4 Various

derivatives of antibodies:

Depending on the clinical

application, antibodies have

been made in different formats

using genetic engineering.

Schematic representation of

scFv comprising the variable

region of heavy and light

chains joined by a peptide

linker, bispecific antibody

capable of binding to two

different targets, antibody-

drug conjugate to deliver the

drug to specific cells, and Fab

fragments is shown

412

S. K. Gupta and P. Chaudhary