The price per patient for protein-based and monoclonal antibody (mAb) therapies runs into thousands of dollars per patient each year. These therapies cost considerably more to manufacture than small molecules. Hence, if mammalian or insect cell lines expressing high protein titres can be selected and optimized for protein expression using microscale bioreactor models early in development, then manufacturing costs can be reduced significantly…
Tag: <span>monoclonal antibody</span>
The purification of PHB-01 plantibody derived from tobacco leaves imposed difficulties when the plantibody solid-liquid extraction design was performed. Thus, our study focused on assessing a combination of an aqueous two-phase extraction (ATPE) procedure and affinity chromatography for solving some of the issues in plantibody purification. This was done using a complete factorial redesign, different polyethylene glycol (PEG)/K2PO4 proportions, and pH values in each partitioning variant.
Cation exchange chromatography is typically utilized in bind-and-elute mode for monoclonal antibody purification. However, during purification process development for a novel monoclonal antibody (MAb) intended for clinical use, it was determined that bind-and-elute conditions were not sufficient for removing significant levels of antibody aggregate. Based on preliminary purification data, an alternative purification method, operation of the cation exchange process in flow-through mode, was investigated.
Rocker bag bioreactors have been used successfully in cultivating cells because they provide good nutrient distribution and cell suspension while eliminating the need to validate cleaning and sterilization. Therefore, this study examined the long-term performance of a 50 L single-use bag bioreactor on a rocking platform in CB.Hep-1 monoclonal antibody (mAb) production. For such a purpose, the bioreactor was operated in a continuous mode with a mixture of serum-free media (SFM) for 62 days, and with protein-free medium (PFM) for another 62 days…
This article reports the average titers and yields currently attained with commercially manufactured biopharmaceuticals expressed by microbial systems such as E. coli and yeasts. A recent BioProcessing Journal article comparably covered results from the first phase of this study concerning historical titers and yields attained for commercial-scale biopharmaceutical production using mammalian cells (e.g., CHO). As with this prior mammalian component, public domain data concerning titers and yields attained with microbially manufactured products were obtained using all available sources.
Antibody-dependent cellular phagocytosis (ADCP), which relies on macrophages to attack and devour tumor cells following antibody binding, is a potentially useful mechanism of action (MOA) for antibody drug developers and vaccine makers to consider in determining product efficacy. Unfortunately, it is often ignored in favor of more accessible MOAs driving biological function such as antibody-dependent cellular cytotoxicity (ADCC) because the assays are tedious to prepare, perform, and reproduce.
The SARS-CoV-2 spike protein S2 subunit plays an essential role in the virus-host cell membrane fusion process. Therefore, the subject of this study was to characterize the gamma-immunoglobulin (IgG) response, in a group of COVID-19 convalescent patients, against the S2 subunit with eight linear peptides to generate a monoclonal antibody (mAb) against the immunodominant linear peptide to be used for therapeutic and diagnostic purposes. Results of antibody percentages against assessed linear peptides were 100% for A21P73, A21P74, A21P75, A21P76, M20P51, M20P65, M20P83, and 66.7% for M20P85. Plasma samples were also used for purifying IgG to corroborate specificity against the same linear peptides, where results reproduced those applying plasmas directly to ELISA-plates. Within these peptides, A21P75 was chosen as immunodominant (100% of recognition with higher absorbance). The A21P75 linear peptide showed poor immunogenicity in mice (1:4000–8000 after four doses), allowing the generation of a CB.HS2A21P75 hybridoma for mAb production that recognized the A21P75 linear peptide with middle-to-high affinity constant (Kaff) (0.8×108 M-1).
This study concludes that the A21P75 linear peptide is the assessed immunodominant linear peptide for this COVID-19 convalescent patient group. This peptide is located in the HR1 region that plays an important role in SARS-CoV-2 host cell membrane fusion process and is highly conserved between SARS-CoV-2 and SARS-CoV. Thus, due to CB.S2A21P75 mAb specificity and Kaff, it might be the proper reagent to study inhibition of virus-host cell membrane fusion, and as a diagnostic reagent for coronavirus. Finally, the combination of A21P75 linear peptide with other peptides (e.g., receptor binding domain [RBD]) could be suitable reagents for the development of vaccines and therapeutic antibodies with virus infection-blocking capacity.
SARS-CoV-2 is an enveloped, positive-strand RNA virus that contains four structural proteins: spike, envelope, membrane, and nucleocapsid (N-protein). The N-protein participates in virus RNA packaging and particle release, is conserved within SARS-CoV-2 isolates, is highly immunogenic, and is abundantly expressed during SARS-CoV-2 infection. For these reasons, the N-protein could be used as a marker for detecting SARS-CoV-2 in early infection when antibodies against SARS-CoV-2 have not been produced yet. This paper describes the production and characterization of mouse monoclonal antibodies (mAb) and rabbit polyclonal antibodies (pAb) specific for the M20P19 peptide (N-protein linear epitope) for detection purposes. For this study, B-cell hybridomas were generated from mice independently immunized with two different M20P19 peptide-carrier protein conjugates: (1)Â meningococcal protein P64K; and (2)Â the keyhole limpet hemocyanin (KLH). Rabbits were also independently immunized with these two immunogens. Study results demonstrated that the M20P19 peptide was very immunogenic in mice and rabbits, and both mAb and pAb specifically recognized the non-conjugated M20P19 peptide, conjugated M20P19 peptide, and N-protein with high affinity and specificity, which could allow SARS-CoV-2 detection by different analytical techniques. This study corroborated that specific and high affinity constant mAb and pAb against the M20P19 peptide can be used as biological reagents for specific and rapid SARS-CoV-2 detection, mainly in tissue samples.