Phage display technology has played a key role in the remarkable progress of discovering and optimizing antibodies for diverse applications, particularly antibody-based drugs. This technology was initially developed by George Smith in the mid-1980s and applied by John McCafferty and Gregory Winter to antibody engineering at the beginning of 1990s. Here, we compare nine phage display antibody libraries published in the last decade, which represent the state of the art in the discovery and development of therapeutic antibodies using phage display. We first discuss the quality of the libraries and the diverse types of antibody repertoires used as substrates to build the libraries, i.e., naïve, synthetic, and semisynthetic. Second, we review the performance of the libraries in terms of the number of positive clones per panning, hit rate, affinity, and developability of the selected antibodies. Finally, we highlight current opportunities and challenges pertaining to phage display platforms and related display technologies.

We describe here the design, construction and validation of ALTHEA Gold Libraries™. These single-chain variable fragment (scFv), semisynthetic libraries are built on synthetic human well-known IGHV and IGKV germline genes combined with natural human complementarity-determining region (CDR)-H3/J_H (H3J) fragments. One IGHV gene provided a universal V_H scaffold and was paired with two IGKV scaffolds to furnish different topographies for binding distinct epitopes. The scaffolds were diversified at positions identified as in contact with antigens in the known antigen-antibody complex structures. The diversification regime consisted of high-usage amino acids found at those positions in human antibody sequences. Functionality, stability and diversity of the libraries were improved throughout a three-step construction process. In a first step, fully synthetic primary libraries were generated by combining the diversified scaffolds with a set of synthetic neutral H3J germline gene fragments. The second step consisted of selecting the primary libraries for enhanced thermostability based on the natural capacity of Protein A to bind the universal V_H scaffold. In the third and final step, the resultant stable synthetic antibody fragments were combined with natural H3J fragments obtained from peripheral blood mononuclear cells of a large pool of 200 donors. Validation of ALTHEA Gold Libraries™ with seven targets yielded specific antibodies in all the cases. Further characterization of the isolated antibodies indicated K_D values as human IgG1 molecules in the single-digit and sub-nM range. The thermal stability (Tm) of all the antigen-binding fragments was 75°C–80°C, demonstrating that ALTHEA Gold Libraries™ are a valuable source of specific, high affinity and highly stable antibodies.

The implementation and validation of anti-SARS-CoV-2 IgG serological assays are reported in this paper. S1 and RBD proteins were used to coat ELISA plates, and several secondary antibodies served as reporters. The assays were initially validated with 50 RT-PCR positive COVID-19 sera, which showed high IgG titers of mainly IgG1 isotype, followed by IgG3. Low or no IgG2 and IgG4 titers were detected. Then, the RBD/IgG assay was further validated with 887 serum samples from RT-PCR positive COVID-19 individuals collected at different times, including 7, 14, 21, and 40 days after the onset of symptoms. Most of the sera were IgG positive at day 40, with seroconversion happening after 14–21 days. A third party conducted an additional performance test of the RBD/IgG assay with 406 sera, including 149 RT-PCR positive COVID-19 samples, 229 RT-PCR negative COVID-19 individuals, and 28 sera from individuals with other viral infections not related to SARS-CoV-2. The sensitivity of the assay was 99.33%, with a specificity of 97.82%. All the sera collected from individuals with infectious diseases other than COVID-19 were negative. Given the robustness of this RBD/IgG assay, it received approval from the sanitary authority in Mexico (COFEPRIS) for production and commercialization under the name UDISTEST-V2G^®.

This study examined the response of osteoblast-like cells to siloxane-based biohybrid materials obtained via the sol–gel process and modified with titania or with titania and calcium. The biohybrids’ ability to form apatite and their degradation process were assessed in vitro via immersion in PBS and SBF. Characterization was performed using XRD, FT-IR, and SEM. The Siloxane-Ti biohybrid exhibited rutile crystalline phase formation, high stability, and resistance to hydrolytic degradation, but did not induce apatite growth. In contrast, the Ca-containing biohybrid showed a vitreous structure with significant surface morphology changes and increased degradability. The Ca-modification enhanced the precipitation of apatite layers, which was attributed to the presence of Ti-OH and Si-OH groups. Immersion of the Siloxane-Ti-Ca biohybrids in PBS produced a flake-like apatite-like mineral structure from the first day of immersion, while immersion in SBF led to a cauliflower-like apatite structure after 28 days. Osteoblast response was evaluated through cell adhesion, proliferation, and differentiation on the hybrid materials. The Siloxane-Ti hybrid promoted osteoblast adhesion and proliferation, whereas the Siloxane-Ti-Ca reduced cell viability and osteonectin expression at 21 days. The Siloxane-Ti hybrid also increased osteonectin expression at early stages, with osteopontin and osteocalcin expression emerging at later stages, indicating progression from early to mature stages stimulated by the hybrid material. The findings suggest that titania enhances the biocompatibility of siloxane materials and shows significant potential for bone tissue regeneration without added growth factors.