Industrial purification of pharmaceutical antibodies: development, operation, and validation of chromatography...

The chromatography technique was first invented in 1901 by Michael S. Tswett, a Russian botanist, to separate plant pigments using organic solvents. Further, in 1941, Archer John Porter Martin and R. L. M. Synge modified the technique by packing silica gel into a column. A mixture of amino acids was then separated on the packed column using chloroform and water mixture as the mobile phase. This was the first report on column chromatography. At present, column chromatography is a widely used...

The stability and compatibility of column material with samples are crucial for efficient purification in chromatographic techniques. Various operating parameters such as pH, temperature, or solvent affect the packing of the column material, thereby determining the purification efficiency. The choice of column material also plays an essential role in deciding the operating parameters and can be modified based on the proteins that need to be purified.
Gel Filtration Chromatography
When the...

Affinity chromatography is a powerful technique extensively utilized for separating and purifying specific biomolecules from complex mixtures. It capitalizes on the highly selective binding between an analyte and its counterpart, such as antibody-antigen interactions. The counterpart is immobilized on the stationary phase, forming an affinity column. The stationary phase typically consists of solid support, such as agarose or porous glass beads, immobilizing the affinity ligand. The mobile...

Upstream processing represents a critical phase in biomanufacturing, wherein biological systems such as microorganisms, mammalian cells, or insect cells are cultivated to produce therapeutic proteins, vaccines, enzymes, or other biologically derived products. This phase encompasses all steps from the selection and genetic manipulation of the production organism to the cultivation of cells in bioreactors under tightly controlled environmental conditions.Host Selection and Genetic OptimizationThe...

Industrial insulin production uses genetically engineered E. coli expressing a proinsulin gene controlled by a tryptophan promoter and containing a methionine linker for later cleavage. The cells also carry ampicillin resistance for selective growth. Seed cultures are stored at −80 °C and production begins by thawing a small amount to inoculate starter cultures, which are progressively scaled to a 50,000-L bioreactor. In the bioreactor, E. coli grow in nutrient-rich media under sterile, tightly...

Downstream processing begins once fermentation is complete and involves a series of steps to recover and purify products such as acids, vitamins, antibiotics, or proteins.Cell HarvestingFor example, for intracellular protein-based products, the first step is harvesting the cells. This is typically achieved using centrifugation or filtration to separate the cells from the liquid phase.Cell Disruption for Intracellular ProductsIf the target product is intracellular, the harvested cells must be...