Developing recombinant antibodies for biomarker detection

• Published in: Cancer biomarkers : section A of Disease markers Vol. 6; no. 5-6; pp. 271 - 279
• Main Authors: Baird, Cheryl L, Fischer, Christopher J, Pefaur, Noah B, Miller, Keith D, Kagan, Jacob, Srivastava, Sudhir, Rodland, Karin D
• Format: Journal Article
• Language: English
• Published: Netherlands 01.01.2010
• Subjects: Antibodies, Monoclonal - biosynthesis; Biomarkers, Tumor - analysis; Biomarkers, Tumor - immunology; Humans; Neoplasms - diagnosis; Neoplasms - immunology; Peptide Library; Proteomics - methods
• ISSN: 1574-0153; 1875-8592
• DOI: 10.3233/CBM-2009-0144

Monoclonal antibodies (mAbs) have an essential role in biomarker validation and diagnostic assays. A barrier to pursuing these applications is the reliance on immunization and hybridomas to produce mAbs, which is time-consuming and may not yield the desired mAb. We recommend a process flow for affinity reagent production that utilizes combinatorial protein display systems (e.g., yeast surface display or phage display) rather than hybridomas. These systems link a selectable phenotype--binding conferred by an antibody fragment--with a means for recovering the encoding gene. Recombinant libraries obtained from immunizations can produce high-affinity antibodies (<10 nM) more quickly than other methods. Non-immune libraries provide an alternate route when immunizations are not possible, or when suitable mAbs are not recovered from an immune library. Directed molecular evolution (DME) is an integral part of optimizing mAbs obtained from combinatorial protein display, but can also be used on hybridoma-derived mAbs. Variants can easily be obtained and screened to increase the affinity of the parent mAb (affinity maturation). We discuss examples where DME has been used to tailor affinity reagents to specific applications. Combinatorial protein display also provides an accessible method for identifying antibody pairs, which are necessary for sandwich-type diagnostic assays.