Biologics Production – Page 9 – BioProcessing Journal

Rapid and Effective Monitoring of Baculovirus Concentrations in Bioprocess Fluid Using the ViroCyt® Virus Counter®

The licensing of recombinant vaccines produced using the baculovirus expression vector system (BEVS) has cleared the way for the production of a variety of biopharmaceuticals produced using this technology. Obtaining accurate estimates of both total and infectious baculovirus titer in upstream and downstream bioprocess fluids is one of many process controls that will need to be addressed during the development phase of a product’s lifecycle. Traditional plaque-titer methods require 5–7 days of incubation in order to reveal plaques that may be enumerated, and is further complicated by plaques created by multiple viruses that may be scored as a single plaque, thereby lowering the titer estimate. Titer assays based on polymerase chain reaction (PCR) have been developed, but they measure the presence of baculovirus genes, not virus particles. This often results in titers one or two logs higher than the actual titer. Immunoassays correlate with host cell infection and virus replication, but they too can be time-consuming and difficult to interpret. Our goal was to identify a method that would provide estimates of both total and infectious virus particles in as close to real-time as possible. We have evaluated the ViroCyt Virus Counter over the course of three years and have found it to provide accurate and reproducible estimates of both titer types in as little as 30 minutes. We have created an algorithm that converts total virus particle counts into estimates of infectious titer and tested these values in virus amplifications. The Virus Counter method of titer determination has also been used to track the quantity of virus particles in the culture supernatant of stirred-tank bioreactors infected with standard baculovirus stocks and with baculovirus-infected insect cells (BIIC)…

Biologics Production

baculovirus express system baculovirus expression system baculovirus titer baculovirus vector BEVS biologics biopharmaceutical manufacturing biopharmaceuticals cell counting cell culture cgmp immunoassays insect cell culture recombinant vaccines vaccine production viral titer virocyt

Optimising Vaccine Process Scale-Up of Attachment-Dependent Cells Using Micro Bioreactors and Microcarriers

The trend for increased vaccine production is being driven by the requirement to produce affordable prophylactic vaccines for use in emerging markets, and also for newer types of therapeutic vaccines to treat an ever-increasing array of diseases such as cancer. These drivers, coupled with the current need to produce vaccines rapidly for pandemic threats and seasonal influenza prevention, has lead to the investigation of rapid method development for optimising the scale-up of cGMP-compliant manufacturing processes…

Biologics Production

attachment-dependent cells biologics biopharmaceutical manufacturing cell culture cgmp microbioreactor microcarriers pandemic threats process scale-up therapeutic vaccines vaccine production

Impact of Process Loading on Optimization and Scale-Up of TFF Microfiltration

Tangential flow filtration (TFF) microfiltration has been used as one of the choices for clarification of mammalian cell or microbial cell culture in the biopharmaceutical industry. Unlike the ultrafiltration process for protein concentration and the diafiltration application where the feed solution is relatively clean (free of colloids or larger particles after the clarification/purification process), the microfiltration process needs to handle a rather high-fouling feed stream such as cells, cell debris, colloids, etc. In a previously published article, we discussed that a TFF microfiltration step is limited by a maximum throughput or capacity obtainable under a given set of operating conditions. Some distinct microfiltration characteristics, such as critical permeate flux, permeate flux control, and maximum throughput were explained in that article…

Biologics Production

biologics biopharmaceutical manufacturing cell culture purification critical permeate flux diafiltration downstream purification maximum throughput microfiltration permeate flux control tangential flow filtration tff ultrafiltration

Insect Cell-Based Recombinant Adeno-Associated Virus Production: Molecular Process Optimization

An increasing number of clinical trials, and the recent approval of the first gene therapy in Europe, alipogene tiparvovec (Glybera®, uniQure), holds promise for recombinant adeno-associated virus (rAAV) to become a mainstay in clinical practice. Since the molecular cloning of AAV in the 1980s, a plethora of production protocols/manufacturing systems for generating rAAV vectors have been developed. uniQure’s manufacturing platform, which has received validation through regulatory approval, is also capable of supporting industrial-scale production based on the baculovirus expression vector system (BEVS) and insect cells. In this paper, we review the molecular process optimization of the various components of uniQure’s rAAV production platform…

Biologics Production

alipogene tiparvovec baculovirus expression systems BEVS glybera insect cells molecular cloning raav recombinant adeno-associated recombinant virus uniqure viral vector

Production of Kojic Acid by Aerobic Aspergillus Fermentation

Kojic acid is produced industrially by the Aspergillus species using aerobic fermentation processes. Kojic acid has applications in several fields such as the pharmaceutical, food production, cosmetics and dermatology, agriculture, and chemical industries. The production of kojic acid is greatly increasing, based on the demands of these industries, and studies focused on improved processes are ongoing. This article will discuss the methods written about by various members of the scientific community…

Biologics Production

aerobic fermentation aspergillus kojic acid microbial fermentation

Validation of a Chromogenic Substrate Method for Biological Activity Quantification of Streptokinase

The use of thrombolytic agent, streptokinase (SK), has been adapted to treat patients with the medical condition, hemorrhoids, in a new pharmaceutical formulation. Part of the development process included studies for measuring SK biological activity in the suppository product. Thus, the main objective in this study was in adapting and validating an established chromogenic substrate method for biological activity quantification of SK extracted from suppositories. By using several solutions and methods described in this paper, results revealed 103.82–124.99% of SK biological activity recovery and high molecular integrity. The chromogenic substrate method was specific for SK, and linear from 200– 600 IU mL-1 (R2 = 0.995 ± 0.002) with a variation coefficient of less than 7.11% and 12.62% for repeatability and reproducibility experiments, respectively. SK biological activity values estimated by the chromogenic substrate method were comparable with those estimated by the clot lysis method used as the reference. We have concluded that the method validated in this study was specific, accurate, and precise for quantifying biological activity of SK extracted from a suppository formulation intended to treat patients with acute hemorrhoids.

Biologics Production

biological activity quantification chromogenic substrate streptokinase thrombolytic agents

Validation Study of the Vi-CELL XR for Dendritic Cell Counting

Dendritic cell (DC) population is a key functional constituent of cell-based immunotherapy drugs. The correct cell count and adequate viability of DCs are one of the quality control criteria for the final product release. The number of viable DCs is historically determined by microscopy using a manual counting method: Bürker chamber, and trypan blue dye for dead cell exclusion. The manual method can have significant variability between cell counts determined by different people performing the procedure, which may contribute to an unstable manufacturing process. The manual method is also timeconsuming for the operator. An automated cell counting process helps remove the variability between operators and can free up the operator for other tasks. The Vi-CELL® XR is an automated cell counting and viability analyzer that uses the trypan blue dye exclusion method. The Vi-CELL was evaluated as a suitable method for quality control of DC counts and viability for a dendritic cell-based biologic drug. The test for Vi-CELL counting accuracy was performed three times each on known concentration control beads, under the same operating conditions. The diameter and circularity of dendritic and lymphocyte cells was determined by a NIKON™ Eclipse microscope to set up the “correct recognition of DC. The size range for DC was established so that lymphocytes could be excluded. The number of total DC, viable and also non-viable, were analyzed and compared to Bürker chamber counting. There was no significant difference between the DC count obtained by Vi-CELL and by Bürker chamber. Vi-CELL automated cell counting was established as a method which is accurate and suitable for use with dendritic cells.

Biologics Production

automated cell counting dc dendritic cells immunotherapy drugs lymphocytes vi-cell xr viability analyzer viable cell counts

Contract Pharma Manufacturer Optimizes Quality Check Process With the Cubis Lab Balance

As a contract manufacturing organization (CMO) in this environment, it is essential for hameln pharma to deliver goods to their customers reliably, in terms of quantity, delivery time, and quality. Therefore, the topic of quality assurance plays a huge role— for both final packaged units as well as for bulk products. After a long-term analysis of the bulk product weighing process, hameln pharma’s objective was to significantly streamline the time-consuming process of manual counting and visual inspection. Moreover, taking cGMP standards into account, they wanted to optimize their existing process in order to remove any possibility of incomplete package volumes. Katrin Strasser, hameln pharma’s operational excellence expert, explained that: (1) packing box weights fluctuated with humidity levels during their manufacture; (2) labels for product packages varied; and (3) the actual number of items in a box carton were not always consistent

Biologics Production

biopharmaceuticals bulk weighing process cgmp cmo contract manufacturing organization lab balance

Failure Mode and Effect Analysis (FMEA) as a Quality by Design (QbD) Tool for Managing Biopharmaceutical Product Development and Manufacturing Risks

This paper examines FMEA as a QbD tool. If FMEA’s strengths are used and weaknesses understood, it can be effectively used within a QbD feedback algorithm to identify, analyze, prioritize, and remediate biopharmaceutical development and manufacturing risks…

Biologics Production

biopharmaceuticals failure mode fmea qbd quality by design

Which Factors to Consider When Selecting an Analytical Method for Cell Culture Fermentation: A Comparison of Four Different Metabolic Analyzer Instruments

Mammalian cell culture processes require an in depth understanding of inputs and outputs in order to maximize productivity, efficiency, and product quality. Daily monitoring of essential metabolites, nutrients, and protein titer using at-line analyzers are the building blocks of drug manufacturing process development and characterization. In order to better understand how these instruments perform, a comparative analysis was conducted using two different classes of metabolicsensing technologies, membrane-based technology (MBT) and absorption photometric-based technology (APBT). Four commercially available instruments were examined using various samples and maintenance conditions in order to emphasize the following testing criteria: specificity, linearity, range, accuracy, and precision. Samples included standard solutions with known metabolite concentrations, cell culture supernatant, and supernatant liquid spiked with additional metabolite solutions. The results from our testing indicated that the photometric analyzers yielded more accurate and consistent results than the membrane-based analyzers and were easier and less time-consuming to maintain. Ultimately, these studies summarized the capabilities and limitations for both types of analyzers and provide a critical summary for instruments used in everyday bioprocess monitoring…

Analytics Biologics Production

adsorption photometry apbt cell culture cell culture fermentation mammalian cell culture matabolic analysers mbt membrane-based technology metabolicsensing technologies

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