Expression of the measles virus proteome by RAPID ELISA for serological assays

Saturday, 14 February 2015
Exhibit Hall (San Jose Convention Center)
Zuena Mushtaq, Center for Personalized Diagnostics, The Biodesign Institute, Arizona State University, Tempe, AZ
Background: Measles virus (MV) infections are the main cause of vaccine-preventable death in children younger than 5 years.  The World Health Organization (WHO) has estimated there are over 20 million cases of measles every year. Currently, diagnostic methods rely on enzyme immunoassays (EIA) to detect IgM or IgG Abs in serum. These commercial assays measure reactivity against the immunodominant N antigen and can have a false negative rates of 20-30%. Centralized testing by clinical labs can delay rapid screening in an outbreak setting. This study aims to develop a rapid molecular diagnostic assay to detect IgM and IgG reactive to five individual MV proteins representing 85% of the measles proteome. Methods: MV genes were subcloned into the Gateway-compatible pANT_cGST vector to generate C-terminal GST fusion proteins. Single MV cistrons were expressed using in vitro transcription/translation (IVTT) with human cell lysate. GST-tagged proteins were captured by anti-GST and measured using a sandwich ELISA for GST expression. Luminescence was detected as relative light units (RLUs). Results: Protein expression of five measles genes of interest, M, N, F, H, and L, was measured. L exhibited the strongest protein expression with an average RLU value of 4.34 x 109.  The average RLU values for M, N, F, and H were 2.89 x 108, 1.91 x 108, 2.10 x 108 and 3.98 x 108, respectively measured.  All proteins were expressed at least 50% greater than control (2.33 x 107 RLU).  Conclusion: These data indicate that MV genes M, N, F, H, and L expressed as recombinant fusion proteins and are suitable for serologic analysis of measles immunity.