Enzyme immunoassay for the detection of anti-Kaposi's sarcoma-associated herpes virus IgG antibodies based on new mosaic protein

Abstract number: 1134_03_316

Kornienko N., Gladysheva M., Ulanova T., Puzyrev V., Burkov A., Loginova L., Obriadina A.

Background:  

Kaposi's sarcoma-associated herpes virus (or human herpes virus type 8 HHV-8) is a recently discovered gamma-herpesvirus associated with 4 clinical and epidemiological variants of Kaposi sarcoma (classic, endemic, iatrogenic, and acquired immunodeficiency virus-associated), primary effusion lymphoma and multicentric Castleman's disease.

Objective:  

The aim of this study was to learn the antigenic properties of new recombinant mosaic protein and to develop and evaluate a screening enzyme immunoassay (EIA) for the detection of anti HHV-8 IgG activity in serum specimens.

Materials and methods:  

Mosaic of 2 antigenic domains from the proteins encoded by open reading frames 65 (140–170 aa) and K8.1 (32–62 aa) of HHV-8 was produced as GST fusion protein to develop an assay for the detection of anti HHV-8 antibodies.

Assay conditions were optimized to reduce the possibility of false positive and false negative results. To validate the specificity and sensitivity of new EIA sera from HIV-infected individuals (n = 163), children (0–15 years) (n = 170), patients with sexual transmitted diseases (STD) (n = 136), and from European normal blood donors (n = 1349) were tested. Serum samples from KS patients (n = 30) were initially tested as positive by immunofluorescence assay with LANA protein. All specimens were additionally tested for IgG anti-HHV-8 activity by commercially available EIA (Vecto HHV-8-IgG-strip,Russia).

Results:  

27 out of 30 HHV-8-positive samples were positive on the novel EIA. Assay sensitivity was calculated at 90%. Coincidence with commercially available EIA was 98.21%. The percentages of positive reactivity in all investigated groups were as follows: 119 for health blood donors, 1.76 for children, 2.21 for STD-patients and 3.38 for HIV-infected. Specificity of the assay was around 98.2%–98.8% and there were no significant differences between health donors/children and groups at highest risk of acquiring HHV8 infection.

Conclusion:  

The artificial mosaic protein used in this study demonstrated significant potential as diagnostic reagent. The new EIA is highly specific diagnostic assay for the detection of anti-HHV-8 activity in serum specimens and may be useful tool for studies of HHV-8 epidemiology.

KSHV seroprevalence in the Russian European population is low both among health donors and HIV-infected people.