Biomaterial Database

Purification of Bacillus anthracis lethal factor by immunosorbent chromatography. 1986

E J Machuga, and G J Calton, and J W Burnett

Lethal factor from B. anthracis (Vollum 1B strain) has been purified 1130-fold by immunosorbent chromatography using a mouse anti-lethal factor monoclonal antibody Sepharose-4B column. The antibody was covalently attached to CNBr activated Sepharose-4B. Lethal factor bound at pH 7 (0.05 M sodium phosphate buffer) and was eluted with buffer containing 4 M NaSCN with 77% recovery of the immunological activity. Pre-elution with 4 M NaCl was effective in eluting non-biospecifically bound proteins. Migration of lethal factor on SDS-polyacrylamide gel electrophoresis indicated a single component (greater than 99% pure) with a molecular weight of 82,000. The effect of a number of dissociation buffers on the antigen-antibody complex has been investigated. Sodium thiocyanate (4 M) in 0.05 M sodium phosphate buffer, pH 7, was the most effective eluting solution, causing complete dissociation of the antigen-antibody complex, while 4M NaI and 4 M NaCl caused 93% and 15% dissociation, respectively. The antigen-antibody complex was found to undergo a reversible dissociation at moderately high pH values. The ionizable group(s) responsible for this dissociation exhibited a pKa value of 9.90. Purified lethal factor exhibited a significant decrease in immunological activity upon freeze-thawing, with 52% loss in potency observed after 3 freeze-thaw cycles.

UI	MeSH Term	Description	Entries
D007163	Immunosorbent Techniques	Techniques for removal by adsorption and subsequent elution of a specific antibody or antigen using an immunosorbent containing the homologous antigen or antibody.	Immunoadsorbent Techniques,Immunoadsorbent Technics,Immunosorbent Technics,Immunoadsorbent Technic,Immunoadsorbent Technique,Immunosorbent Technic,Immunosorbent Technique,Technic, Immunoadsorbent,Technic, Immunosorbent,Technics, Immunoadsorbent,Technics, Immunosorbent,Technique, Immunoadsorbent,Technique, Immunosorbent,Techniques, Immunoadsorbent,Techniques, Immunosorbent
D008807	Mice, Inbred BALB C	An inbred strain of mouse that is widely used in IMMUNOLOGY studies and cancer research.	BALB C Mice, Inbred,BALB C Mouse, Inbred,Inbred BALB C Mice,Inbred BALB C Mouse,Mice, BALB C,Mouse, BALB C,Mouse, Inbred BALB C,BALB C Mice,BALB C Mouse
D004591	Electrophoresis, Polyacrylamide Gel	Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.	Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D004797	Enzyme-Linked Immunosorbent Assay	An immunoassay utilizing an antibody labeled with an enzyme marker such as horseradish peroxidase. While either the enzyme or the antibody is bound to an immunosorbent substrate, they both retain their biologic activity; the change in enzyme activity as a result of the enzyme-antibody-antigen reaction is proportional to the concentration of the antigen and can be measured spectrophotometrically or with the naked eye. Many variations of the method have been developed.	ELISA,Assay, Enzyme-Linked Immunosorbent,Assays, Enzyme-Linked Immunosorbent,Enzyme Linked Immunosorbent Assay,Enzyme-Linked Immunosorbent Assays,Immunosorbent Assay, Enzyme-Linked,Immunosorbent Assays, Enzyme-Linked
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia
D000942	Antigens, Bacterial	Substances elaborated by bacteria that have antigenic activity.	Bacterial Antigen,Bacterial Antigens,Antigen, Bacterial
D001408	Bacillus anthracis	A species of bacteria that causes ANTHRAX in humans and animals.
D001427	Bacterial Toxins	Toxic substances formed in or elaborated by bacteria; they are usually proteins with high molecular weight and antigenicity; some are used as antibiotics and some to skin test for the presence of or susceptibility to certain diseases.	Bacterial Toxin,Toxins, Bacterial,Toxin, Bacterial
D051379	Mice	The common name for the genus Mus.	Mice, House,Mus,Mus musculus,Mice, Laboratory,Mouse,Mouse, House,Mouse, Laboratory,Mouse, Swiss,Mus domesticus,Mus musculus domesticus,Swiss Mice,House Mice,House Mouse,Laboratory Mice,Laboratory Mouse,Mice, Swiss,Swiss Mouse,domesticus, Mus musculus