scholarly journals Process development and scale-up optimization of the SARS-CoV-2 receptor binding domain–based vaccine candidate, RBD219-N1C1

2021 ◽  
Author(s):  
Jungsoon Lee ◽  
Zhuyun Liu ◽  
Wen-Hsiang Chen ◽  
Junfei Wei ◽  
Rakhi Kundu ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Process Development ◽  
Low Cost ◽  
Scale Up ◽  
Neutralizing Antibody ◽  
Protein Antigen ◽  
Vaccine Antigen ◽  
Yeast Fermentation ◽  
Purification Process ◽  
Purification Methods

Abstract A SARS-CoV-2 RBD219-N1C1 (RBD219-N1C1) recombinant protein antigen formulated on Alhydrogel® has recently been shown to elicit a robust neutralizing antibody response against SARS-CoV-2 pseudovirus in mice. The antigen has been produced under current good manufacturing practices (cGMPs) and is now in clinical testing. Here, we report on process development and scale-up optimization for upstream fermentation and downstream purification of the antigen. This includes production at the 1-L and 5-L scales in the yeast, Pichia pastoris, and the comparison of three different chromatographic purification methods. This culminated in the selection of a process to produce RBD219-N1C1 with a yield of >400 mg per liter of fermentation with >92% purity and >39% target product recovery after purification. In addition, we show the results from analytical studies, including SEC-HPLC, DLS, and an ACE2 receptor binding assay that were performed to characterize the purified proteins to select the best purification process. Finally, we propose an optimized upstream fermentation and downstream purification process that generates quality RBD219-N1C1 protein antigen and is fully scalable at a low cost. Key points • Yeast fermentation conditions for a recombinant COVID-19 vaccine were determined. • Three purification protocols for a COVID-19 vaccine antigen were compared. • Reproducibility of a scalable, low-cost process for a COVID-19 vaccine was shown. Graphical abstract

scholarly journals Process Development and Scale-up Optimization of the SARS-CoV-2 Receptor Binding Domain-Based Vaccine Candidate, RBD219-N1C1

2020 ◽  
Author(s):  
Jungsoon Lee ◽  
Zhuyun Liu ◽  
Wen-Hsiang Chen ◽  
Junfei Wei ◽  
Rakhi Kundu ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Process Development ◽  
Low Cost ◽  
Scale Up ◽  
Neutralizing Antibody ◽  
Protein Antigen ◽  
Purification Process ◽  
Good Manufacturing Practices ◽  
Purification Methods ◽  
Selection Of

AbstractA SARS-CoV-2 RBD219-N1C1 (RBD219-N1C1) recombinant protein antigen formulated on Alhydrogel® has recently been shown to elicit a robust neutralizing antibody response against SARS-CoV-2 pseudovirus in mice. The antigen has been produced under current good manufacturing practices (cGMP) and is now in clinical testing. Here, we report on process development and scale-up optimization for upstream fermentation and downstream purification of the antigen. This includes production at the 1 and 5 L scale in the yeast, Pichia pastoris, and the comparison of three different chromatographic purification methods. This culminated in the selection of a process to produce RBD219-N1C1 with a yield of >400 mg per liter of fermentation with >92% purity and >39% target product recovery after purification. In addition, we show the results from analytical studies, including SEC-HPLC, DLS, and an ACE2 receptor binding assay that were performed to characterize the purified proteins to select the best purification process. Finally, we propose an optimized upstream fermentation and downstream purification process that generates quality RBD219-N1C1 protein antigen and is fully scalable at a low cost.

Image Processing for Solar Cell Analysis, Diagnostics and Quality Assurance Inspection

2013 ◽  
pp. 338-375
Author(s):  
Michael G. Mauk
Keyword(s):  
Image Processing ◽  
Image Analysis ◽  
Quality Assurance ◽  
Solar Cell ◽  
Process Development ◽  
Low Cost ◽  
Scale Up ◽  
Image Capturing ◽  
Infrared Cameras ◽  
Primary Focus

Image capturing, processing, and analysis have numerous uses in solar cell research, device and process development and characterization, process control, and quality assurance and inspection. Solar cell image processing is expanding due to the increasing performance (resolution, sensitivity, spectral range) and low-cost of commercial CCD and infrared cameras. Methods and applications are discussed, with primary focus on monocrystalline and polycrystalline silicon solar cells using visible and infrared (thermography) wavelengths. The most prominent applications relate to mapping of minority carrier lifetime, shunts, and defects in solar cell wafers, in various stages of the manufacturing process. Other applications include measurements of surface texture and reflectivity, surface cleanliness, integrity of metallization lines, uniformity of coatings, and crystallographic texture and grain size. Image processing offers the capability to assess large-areas (> 100 cm2) with a non-contact, fast (~ 1 second), and modest cost. The challenge is to quantify and interpret the image data in order to better inform device design, process engineering, and quality control. Many promising solar cell technologies fail in the transition from laboratory to factory due to issues related to scale-up in area and manufacturing throughput. Image analysis provides an effective method to assess areal uniformity, device-to-device reproducibility, and defect densities. More integration of image analysis from research devices to field testing of modules will continue as the photovoltaics industry matures.

scholarly journals Rapid development of SARS-CoV-2 receptor binding domain-conjugated nanoparticle vaccine candidate

2020 ◽  
Author(s):  
Yin-Feng Kang ◽  
Cong Sun ◽  
Zhen Zhuang ◽  
Run-Yu Yuan ◽  
Qing-Bing Zheng ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Vaccine Development ◽  
Vaccine Candidate ◽  
Rapid Development ◽  
Scale Up ◽  
Physical Stability ◽  
Covalent Bond ◽  
Neutralizing Antibody ◽  
Economic Losses

AbstractThe ongoing of coronavirus disease 2019 (COVID-19) pandemic caused by novel SARS-CoV-2 coronavirus, resulting in economic losses and seriously threating the human health in worldwide, highlighting the urgent need of a stabilized, easily produced and effective preventive vaccine. The SARS-COV-2 spike protein receptor binding region (RBD) plays an important role in the process of viral binding receptor angiotensin-converting enzyme 2 (ACE2) and membrane fusion, making it an ideal target for vaccine development. In this study, we designed three different RBD-conjugated nanoparticles vaccine candidates, RBD-Ferritin (24-mer), RBD-mi3 (60-mer) and RBD-I53-50 (120-mer), with the application of covalent bond linking by SpyTag-SpyCatcher system. It was demonstrated that the neutralizing capability of sera from mice immunized with three RBD-conjugated nanoparticles adjuvanted with AddaVax or Sigma Systerm Adjuvant (SAS) after each immunization was ~8-to 120-fold greater than monomeric RBD group in SARS-CoV-2 pseudovirus and authentic virus neutralization assay. Most importantly, sera from RBD-conjugated NPs groups more efficiently blocked the binding of RBD to ACE2 or neutralizing antibody in vitro, a further proof of promising immunization effect. Besides, high physical stability and flexibility in assembly consolidated the benefit for rapid scale-up production of vaccine. These results supported that our designed SARS-CoV-2 RBD-conjugated nanoparticle was competitive vaccine candidate and the carrier nanoparticles could be adopted as universal platform for future vaccine development.

scholarly journals Protection of Rabbits against Challenge with Rabbit Papillomaviruses by Immunization with the N Terminus of Human Papillomavirus Type 16 Minor Capsid Antigen L2

2007 ◽  
Vol 81 (21) ◽  
pp. 11585-11592 ◽  
Author(s):  
Ratish Gambhira ◽  
Subhashini Jagu ◽  
Balasubramanyam Karanam ◽  
Patti E. Gravitt ◽  
Timothy D. Culp ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Human Papillomavirus ◽  
Genomic Dna ◽  
Low Cost ◽  
Serum Antibody ◽  
Neutralizing Antibody ◽  
Vaccine Antigen ◽  
Small Subset ◽  
Bovine Papillomavirus ◽  
Degree Of Protection

ABSTRACT Current L1 virus-like particle (VLP) vaccines provide type-restricted protection against a small subset of the human papillomavirus (HPV) genotypes associated with cervical cancer, necessitating continued cytologic screening of vaccinees. Cervical cancer is most problematic in countries that lack the resources for screening or highly multivalent HPV VLP vaccines, suggesting the need for a low-cost, broadly protective vaccinogen. Here, N-terminal L2 polypeptides comprising residues 1 to 88 or 11 to 200 derived from HPV16, bovine papillomavirus type 1 (BPV1), or cottontail rabbit papillomavirus (CRPV) were produced in bacteria. Rabbits were immunized with these N-terminal L2 polypeptides and concurrently challenged with CRPV and rabbit oral papillomavirus (ROPV). Vaccination with either N-terminal L2 polypeptides of CRPV effectively protected rabbits from CRPV challenge but not from papillomas induced by cutaneous challenge with CRPV genomic DNA. Furthermore, papillomas induced by CRPV genomic DNA deficient for L2 expression grew at the same rate as those induced by wild-type CRPV genomic DNA, further suggesting that the L2 polypeptide vaccines lack therapeutic activity. Neutralizing serum antibody titers of >15 correlated with protection (P < 0.001), a finding consistent with neutralizing antibody-mediated protection. Surprisingly, a remarkable degree of protection against heterologous papillomavirus types was observed after vaccination with N-terminal L2 polypeptides. Notably, vaccination with HPV16 L2 11-200 protected against cutaneous and mucosal challenge with CRPV and ROPV, respectively, papillomaviruses that are evolutionarily divergent from HPV16. Further, vaccination with HPV16 L2 11-200 generates broadly cross-neutralizing serum antibody, suggesting the potential of L2 as a second-generation preventive HPV vaccine antigen.

Accelerating Laboratory Thin Film Photovoltaic Innovation into Commercial Production

2009 ◽  
Vol 1165 ◽  
Author(s):  
Fred H. Seymour
Keyword(s):  
Thin Film ◽  
Alternative Energy ◽  
Process Development ◽  
Low Cost ◽  
Scale Up ◽  
High Volume ◽  
Commercial Production ◽  
Laboratory Research ◽  
Small Scale ◽  
Managing Risk

AbstractAlternative energy sources such as thin film photovoltaics can be accelerated by improving the rapid and successful transition from laboratory research innovation to commercial production. Most laboratory research and development is on a small scale and its production is in small volumes. It focuses on exploration, discovery, and understanding. When the successful innovation is commercialized, both the scale and the volume increase dramatically and the focus shifts to performance, reliability, yield and cost. This transformation can be accelerated by closely managing risk and by integrating the equipment design and the process development. Also, the cadmium telluride photovoltaic technology has properties that make it more amenable to rapid scale up to low cost and high volume manufacturing.

Image Processing for Solar Cell Analysis, Diagnostics and Quality Assurance Inspection

2013 ◽  
pp. 1426-1462
Author(s):  
Michael G. Mauk
Keyword(s):  
Image Processing ◽  
Image Analysis ◽  
Quality Assurance ◽  
Solar Cell ◽  
Process Development ◽  
Low Cost ◽  
Scale Up ◽  
Image Capturing ◽  
Infrared Cameras ◽  
Primary Focus

Image capturing, processing, and analysis have numerous uses in solar cell research, device and process development and characterization, process control, and quality assurance and inspection. Solar cell image processing is expanding due to the increasing performance (resolution, sensitivity, spectral range) and low-cost of commercial CCD and infrared cameras. Methods and applications are discussed, with primary focus on monocrystalline and polycrystalline silicon solar cells using visible and infrared (thermography) wavelengths. The most prominent applications relate to mapping of minority carrier lifetime, shunts, and defects in solar cell wafers, in various stages of the manufacturing process. Other applications include measurements of surface texture and reflectivity, surface cleanliness, integrity of metallization lines, uniformity of coatings, and crystallographic texture and grain size. Image processing offers the capability to assess large-areas (> 100 cm2) with a non-contact, fast (~ 1 second), and modest cost. The challenge is to quantify and interpret the image data in order to better inform device design, process engineering, and quality control. Many promising solar cell technologies fail in the transition from laboratory to factory due to issues related to scale-up in area and manufacturing throughput. Image analysis provides an effective method to assess areal uniformity, device-to-device reproducibility, and defect densities. More integration of image analysis from research devices to field testing of modules will continue as the photovoltaics industry matures.

scholarly journals Scalable, methanol-free manufacturing of the SARS-CoV-2 receptor binding domain in engineered Komagataella phaffii

2021 ◽  
Author(s):  
Neil C Dalvie ◽  
Andrew M Biedermann ◽  
Sergio A Rodriguez-Aponte ◽  
Christopher A Naranjo ◽  
Harish D Rao ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Large Scale ◽  
Scale Up ◽  
Binding Domain ◽  
Vaccine Antigen ◽  
Scale Production ◽  
Receptor Binding Domain ◽  
Methanol Induction ◽  
Vaccine Candidates ◽  
Komagataella Phaffii

Prevention of COVID-19 on a global scale will require the continued development of high-volume, low-cost platforms for the manufacturing of vaccines to supply on-going demand. Vaccine candidates based on recombinant protein subunits remain important because they can be manufactured at low costs in existing large-scale production facilities that use microbial hosts like Komagataella phaffii (Pichia pastoris). Here, we report an improved and scalable manufacturing approach for the SARS-CoV-2 spike protein receptor binding domain (RBD); this protein is a key antigen for several reported vaccine candidates. We genetically engineered a manufacturing strain of K. phaffii to obviate the requirement for methanol-induction of the recombinant gene. Methanol-free production improved the secreted titer of the RBD protein by >5x by alleviating protein folding stress. Removal of methanol from the production process enabled scale up to a 1,200 L pre-existing production facility. This engineered strain is now used to produce an RBD-based vaccine antigen that is currently in clinical trials and could be used to produce other variants of RBD as needed for future vaccines.

scholarly journals SARS-CoV-2 RBD219-N1C1: A Yeast-Expressed SARS-CoV-2 Recombinant Receptor-Binding Domain Candidate Vaccine Stimulates Virus Neutralizing Antibodies and T-cell Immunity in Mice

2020 ◽  
Author(s):  
Jeroen Pollet ◽  
Wen-Hsiang Chen ◽  
Leroy Versteeg ◽  
Brian Keegan ◽  
Bin Zhan ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Low Cost ◽  
Neutralizing Antibody ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Wild Type ◽  
Cell Immunity

AbstractThere is an urgent need for an accessible and low-cost COVID-19 vaccine suitable for low- and middle-income countries. Here we report on the development of a SARS-CoV-2 receptor-binding domain (RBD) protein, expressed at high levels in yeast (Pichia pastoris), as a suitable vaccine candidate against COVID-19. After introducing two modifications into the wild-type RBD gene to reduce yeast-derived hyperglycosylation and improve stability during protein expression, we show that the recombinant protein, RBD219-N1C1, is equivalent to the wild-type RBD recombinant protein (RBD219-WT) in an in vitro ACE-2 binding assay. Immunogenicity studies of RBD219-N1C1 and RBD219-WT proteins formulated with Alhydrogel® were conducted in mice, and, after two doses, both the RBD219-WT and RBD219-N1C1 vaccines induced high levels of binding IgG antibodies. Using a SARS-CoV-2 pseudovirus, we further showed that sera obtained after a two-dose immunization schedule of the vaccines were sufficient to elicit strong neutralizing antibody titers in the 1:1,000 to 1:10,000 range, for both antigens tested. The vaccines induced IFN-γ, IL-6, and IL-10 secretion, among other cytokines. Overall, these data suggest that the RBD219-N1C1 recombinant protein, produced in yeast, is suitable for further evaluation as a human COVID-19 vaccine, in particular, in an Alhydrogel® containing formulation and possibly in combination with other immunostimulants.

Constructive Optimisation of the Propeller Type Mixing Devices Using the Virtual and Rapid Prototyping

2017 ◽  
Vol 68 (3) ◽  
pp. 453-458 ◽  
Author(s):  
Daniel Besnea ◽  
Alina Spanu ◽  
Iuliana Marlena Prodea ◽  
Gheorghita Tomescu ◽  
Iolanda Constanta Panait
Keyword(s):  
Rapid Prototyping ◽  
Low Cost ◽  
Scale Up ◽  
Three Dimensional ◽  
Rapid Identification ◽  
Multidisciplinary Optimization ◽  
External Diameter ◽  
Process Industries ◽  
Parametric Cad ◽  
Shaft Torque

The paper points out the advantages of rapid prototyping for improving the performances/constructive optimization of mixing devices used in process industries, here exemplified to propeller types ones. The multidisciplinary optimization of the propeller profile affords its design using parametric CAD methods. Starting from the mathematical curve equations proposed for the blade profile, it was determined its three-dimensional virtual model. The challenge has been focused on the variation of propeller pitch and external diameter. Three dimensional ranges were manufactured using the additive manufacturing process with Marker Boot 3D printer. The mixing performances were tested on the mixing equipment measuring the minimum rotational speed and the correspondent shaft torque for complete suspension achieved for each of the three models. The virtual and rapid prototyping method is newly proposed by the authors to obtain the basic data for scale up of the mixing systems, in the case of flexible production (of low quantities), in which both the nature and concentration of the constituents in the final product varies often. It is an efficient and low cost method for the rapid identification of the optimal mixing device configuration, which contributes to the costs reduction and to the growing of the output.

scholarly journals A SARS-CoV-2 neutralizing antibody with extensive Spike binding coverage and modified for optimal therapeutic outcomes

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yu Guo ◽  
Lisu Huang ◽  
Guangshun Zhang ◽  
Yanfeng Yao ◽  
He Zhou ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Complex Structure ◽  
Neutralizing Antibody ◽  
Economic Consequences ◽  
Binding Motif ◽  
Global Public Health ◽  
Health Crisis ◽  
Public Health Crisis ◽  
Monkey Model ◽  
Therapeutic Outcomes

AbstractCOVID-19 pandemic caused by SARS-CoV-2 constitutes a global public health crisis with enormous economic consequences. Monoclonal antibodies against SARS-CoV-2 can provide an important treatment option to fight COVID-19, especially for the most vulnerable populations. In this work, potent antibodies binding to SARS-CoV-2 Spike protein were identified from COVID-19 convalescent patients. Among them, P4A1 interacts directly with and covers majority of the Receptor Binding Motif of the Spike Receptor-Binding Domain, shown by high-resolution complex structure analysis. We further demonstrate the binding and neutralizing activities of P4A1 against wild type and mutant Spike proteins or pseudoviruses. P4A1 was subsequently engineered to reduce the potential risk for Antibody-Dependent Enhancement of infection and to extend its half-life. The engineered antibody exhibits an optimized pharmacokinetic and safety profile, and it results in complete viral clearance in a rhesus monkey model of COVID-19 following a single injection. These data suggest its potential against SARS-CoV-2 related diseases.

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