Second generation of pseudotype-based serum neutralization assay for Nipah virus antibodies: Sensitive and high-throughput analysis utilizing secreted alkaline phosphatase

► A novel serum neutralization test for Nipah virus (NiV) infection was developed. ► This assay employs SEAP-expressing VSV pseudotype bearing NiV-F/G proteins. ► It is performed by measuring SEAP in supernatant using an ELISA plate reader. ► Various species sera were tested and showed identical serostatus to live NiV test. ► The assay is useful for high-throughput screening for NiV neutralizing antibodies.