A Novel Peptide ELISA for Universal Detection of Antibodies to Human H5N1 Influenza Viruses

Abstract

BackgroundActive serologic surveillance of H5N1 highly pathogenic avian influenza (HPAI) virus in humans and poultry is critical to control this disease. However, the need for a robust, sensitive and specific serologic test for the rapid detection of antibodies to H5N1 viruses has not been met.Methodology/Principal FindingsPreviously, we reported a universal epitope (CNTKCQTP) in H5 hemagglutinin (HA) that is 100% conserved in H5N1 human isolates and 96.9% in avian isolates. Here, we describe a peptide ELISA to detect antibodies to H5N1 virus by using synthetic peptide that comprises the amino acid sequence of this highly conserved and antigenic epitope as the capture antigen. The sensitivity and specificity of the peptide ELISA were evaluated using experimental chicken antisera to H5N1 viruses from divergent clades and other subtype influenza viruses, as well as human serum samples from patients infected with H5N1 or seasonal influenza viruses. The peptide ELISA results were compared with hemagglutinin inhibition (HI), and immunofluorescence assay and immunodot blot that utilize recombinant HA1 as the capture antigen. The peptide ELISA detected antibodies to H5N1 in immunized animals or convalescent human sera whereas some degree of cross-reactivity was observed in HI, immunofluorescence assay and immunodot blot. Antibodies to other influenza subtypes tested negative in the peptide-ELISA.Conclusion/SignificanceThe peptide-ELISA based on the highly conserved and antigenic H5 epitope (CNTKCQTP) provides sensitive and highly specific detection of antibodies to H5N1 influenza viruses. This study highlighted the use of synthetic peptide as a capture antigen in rapid detection of antibodies to H5N1 in human and animal sera that is robust, simple and cost effective and is particularly beneficial for developing countries and rural areas.

The Development of a Universally Conserved M2E and Hemagglutinin Peptide-based ELISA Method Against Influenza A

“…However, these studies used whole M2e protein rather than the more conserved N-terminal 24-amino-acid domain of the protein to test for antibodies. Specific peptide-based ELISA methods have several advantages over whole protein-based methods, such as high specificity and, potentially, a simple, rapid, sensitive, and relatively inexpensive diagnostic kit [6]. In this study, positive responses against the M2e and H3 peptide sequences were seen.…”

“…Serological techniques still play a major role in influenza surveillance, vaccine development and evaluation, and diagnosis [5]. The Hemagglutinin Inhibition (HI) serological test is considered the "gold standard" for subtyping but this method is limited by low sensitivity and subtype cross-reactivity [6]. Influenza A viruses have structures that easily change conformation.…”

Application of Peptide for Improving Animal Health and Livestock Productivity

“…Oleh karena itu, vaksin peptida yang berasal dari bagian antigenik virus dapat menjadi solusi. Wang et al (2002) (Velumani et al 2011).…”

Uso de un péptido sintético de la glucoproteína 36 del VIH 2 en mezclas de antígenos para el inmunodiagnóstico de la infección por VIH tipo 1 y 2

“…Entre las más empleadas se encuentran técnicas inmunoenzimáticas de tercera y cuarta generación, como la quimioluminiscencia (ChLIA) y los ELISA 2 . Los antígenos usados en el recubrimiento de los ELISA pueden ser producto de lisados virales, proteínas recombinantes y péptidos sintéticos, siendo estos últimos ampliamente utilizados en la actualidad, en busca de mayor sensibilidad y especificidad 3 UMELISA HIV 1 + 2 RECOMBINANT gp120rec (0,5 g/ml) + gp41rec (1 g/ml) + p24rec (1 g/ml) + gp36rec (0,5 g/ml)+ p24s V-1 gp120rec (0,5 g/ml) + gp41rec (1 g/ml) + p24rec (1 g/ml) + gp36 (5)s (1 g/ml) V-2 gp120rec (0,5 g/ml) + gp41rec (1 g/ml) + p24rec (1 g/ml) + gp36 (5)s (2 g/ml) V-3 gp120rec (0,5 g/ml) + gp41rec (1 g/ml) + p24rec (1 g/ml) + gp36 (5)s (3 g/ml) V-4 gp120rec (0,5 g/ml) + gp41rec (1 g/ml) + p24rec (1 g/ml) + gp36 (5)s (4 g/ml) gp: glucoproteína; p: proteína; rec: recombinante; s: sintético.…”