Quantification of an Epstein-Barr virus-associated membrane antigen component

Area of Science:

• Biochemistry
• Immunology
• Cell Biology

Background:

• Membrane antigen (MA) gp340 is a component of B95-8 cell membranes.
• Previous methods lacked efficient ways to isolate and quantify gp340.

Purpose of the Study:

• To develop a method for preparing purified, radiolabeled gp340.
• To establish a radioimmunoassay for quantifying gp340.
• To investigate gp340 expression levels.

Main Methods:

• Utilized sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) for gp340 isolation.
• Employed urea-assisted renaturation to maintain antigenicity of [125I]gp340.
• Developed a radioimmunoassay for antigen quantification.

Main Results:

• Achieved good yields of antigenic [125I]gp340.
• Established a sensitive radioimmunoassay for gp340 quantification.
• Demonstrated that cell membrane MA is a superior source for large-scale gp340 production compared to Epstein-Barr virus envelope.
• Observed increased gp340 expression upon 12-O-tetradecanoyl-phorbol-13-acetate (TPA) treatment.

Conclusions:

• A robust method for preparing and radiolabeling gp340 was established.
• The developed radioimmunoassay allows for the quantification of gp340.
• Cell membrane-derived gp340 is suitable for large-scale applications.
• TPA treatment significantly upregulates gp340 expression in cells.