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Use of avidity reagent

Use of avidity reagent. Panbio Buffered Avidity Reagent. Mild protein-denaturing solution that may be useful in differentiating recent infections from past infections , particularly during a prolonged IgM response.

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Use of avidity reagent

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  1. Use of avidity reagent

  2. Panbio Buffered Avidity Reagent • Mild protein-denaturing solution that may be useful in differentiating recent infections from past infections, particularly during a prolonged IgM response. • May be particularly useful in WNV infection as IgM antibody has been shown to persist for >500 days in approximately 60% of cases. • May remove low avidity antibodies thus resulting in a change in the IgG absorbance level. This change is indicative of the avidity of the antibodies.

  3. Panbio Buffered Avidity Reagent • Low avidity IgG antibodies = early stage of infection (removed by avidity reagent) • High avidity IgG antibodies = past infection. • To determine the Avidity Index of a patient sample, perform the Panbio WNV IgG ELISA with and without Buffered Avidity Reagent. • The following slide illustrates where to incorporate the Buffered Avidity Reagent.

  4. Avidity Reagent procedure

  5. Calculation Absorbance B X 100 Avidity index (%) = Absorbance A Absorbance A = Standard Panbio WNV IgG ELISA result Absorbance B = Panbio WNV IgG ELISA result with avidity reagent

  6. Interpretation of results • The interpretation of avidity results varies depending on disease state, concentration of urea and ELISA used. It is recommended that laboratories perform in-house validation by comparing acute and convalescent samples to determine an acceptable cut-off for that disease state with the Panbio Buffered Avidity Reagent before interpretation of test sample results. • Inhouse studies have found that 40 days after infection with WNV patient have an Avidity Index of >40% (Fox et al., 2005)

  7. Fox et al (2005): Summary • Fox, J.L. et al. (2005) Immunoglobulin G Avidity in the Differentiation Between Early and Late Antibody Response to West Nile Virus. Poster presented at the 2005 Clearwater Virology Meeting, USA. • Thirteen seroconversion panels, each consisting of between four and seven plasma samples collected from U.S. blood donors classified as WNV RNA positive by nucleic acid technology (NAT) screening, were obtained. The index sample (Day 0) was positive by a WNV Transcription Mediated Amplication (TMA) assay and non-reactive when tested for WNV IgM. Each panel contained at least two follow up specimens taken at various intervals ranging from 10 to 208 days post index. All samples were independently tested by the CDC using PRNT. • Samples were tested on the Panbio WNV IgM ELISA and the IgG ELISA with and without avidity reagent.

  8. Fox et al (2005): Results • IgG avidity profiles of "primary" individuals had low Avidity Index (< 40%) for the first 20-30 days post index. • After ~ 40 days post index in these panels, the Avidity Index rose to > 40% (Fig 1). • For the "secondary" samples all positive IgG samples had an Avidity Index of > 55% regardless of the days since the index donation (Fig 1).

  9. Fox et al (2005): Conclusion • When both IgM and IgG are present in a patient sample, IgG avidity testing in conjunction with the WNV ELISA appears to be useful in identifying recent, primary infections.

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