Tag: <span>facs</span>

Large-Scale Growth of Mouse P815 Cells Expressing a Bovine Non-Classical Major Histocompatibility Complex Class I Protein Utilizing a Pitched-Blade Bioreactor

In an attempt to express and isolate BoLA-NC3*00101, a bovine, non-classical, major histocompatibility complex (MHC) class I protein, the mouse mastocytoma cell line P815 was transfected with the NC3*00101 transgene. The transfected cells were checked for expression by flow cytometry and positive stable transfectants were sorted using a fluorescence-activated cell sorter (FACS). A large amount of purified protein is required for the generation of monoclonal antibodies in order to immunize the mice and then boost the host for several months to get highly-specific antibodies. To avoid using a large number of cell culture flasks and shorten the time required to obtain a sufficient quantity of protein, stably-transfected BoLA-NC3*00101-expressing cells were grown in a pitched-blade bioreactor. One week of culturing in the bioreactor yielded a large cell mass which was used to isolate and purify the protein…

Read MoreLarge-Scale Growth of Mouse P815 Cells Expressing a Bovine Non-Classical Major Histocompatibility Complex Class I Protein Utilizing a Pitched-Blade Bioreactor

Biologics Production

biologics class 1 protein facs fluorescence-activated cell sorter histocompatibility hybridoma culture mouse p815 cells pitched-blade bioreactor surface glycoprotein transgene

Statistical Verification that One Round of Fluorescence-Activated Cell Sorting (FACS) Can Effectively Generate a Clonally-Derived Cell Line

Mammalian cell line generation is foundational to protein-based therapeutic product development. Ensuring production cell line “clonality” (i.e., clonal derivation of the production cell line) is a requirement for product registration. Many single-cell deposition approaches have been developed since the inception of traditional limiting-dilution. However, regulatory expectations, in line with the capability of this original cloning technique, still lead many institutions to either execute multiple rounds of single-cell plating or conduct extensive screening immediately post-cloning to demonstrate the single-cell origin of a production cell line. Using fluorescence-activated cell sorting (FACS), we have developed a strategy to verify single-cell deposition that, when combined with statistical analysis, creates a highly-effective and efficient process for generating clonally-derived production cell lines in just one round of plating…

Read MoreStatistical Verification that One Round of Fluorescence-Activated Cell Sorting (FACS) Can Effectively Generate a Clonally-Derived Cell Line

Analytics Biologics Production

clonality cloning techniques facs fluorescence-activated cell mammalian cells protein-based therapeutics statistical analysis