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Clinical and Vaccine Immunology, May 2009, p. 621-627, Vol. 16, No. 5
1071-412X/09/$08.00+0     doi:10.1128/CVI.00474-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Four-Antigen Mixture Containing V-Cyclin for Serological Screening of Human Herpesvirus 8 Infection{triangledown}

Peter D. Burbelo,1* Hannah P. Leahy,1 Sandra Groot,1 Lisa R. Bishop,3 Wendell Miley,2 Michael J. Iadarola,1 Denise Whitby,2 and Joseph A. Kovacs3

Neurobiology and Pain Therapeutics Section, Laboratory of Sensory Biology, National Institute of Dental and Craniofacial Research, Bethesda, Maryland 20892-4410,1 Viral Oncology Section, AIDS and Cancer Virus Program, Science Applications International Corporation-Frederick, NCI-Frederick, Frederick, Maryland,2 Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland 208923

Received 5 December 2008/ Returned for modification 13 January 2009/ Accepted 21 February 2009

Improved diagnostic reagents and testing are currently needed for the serological detection of human herpesvirus 8 (HHV-8) infections. We evaluated the luciferase immunoprecipitation systems (LIPS) for profiling antibody responses to a panel of HHV-8 proteins for diagnosis of Kaposi sarcoma (KS)-infected individuals. Using a pilot serum set, LIPS detected robust antibody responses to several known antigens, and a screen of 14 additional HHV-8 proteins identified v-cyclin as a potentially new diagnostic antigen. In evaluating a training-serum set, a four-antigen panel (K8.1, v-cyclin, ORF65, and a LANA fragment) was found to provide sufficient information for diagnosis. Analysis of a validation serum set using the combined results from these four separate antigen tests showed 100% sensitivity and 100% specificity. Furthermore, a LIPS format using a mixture of the four antigens, which simplifies data collection and analysis, closely matched the diagnostic performance of the combined separate tests (R = 0.95). This four-antigen mixture format analyzed with the validation serum set also showed 100% sensitivity and 100% specificity but was not statistically different from two separate enzyme-linked immunosorbent assays (94% sensitivity and 100% specificity) using baculovirus-produced LANA and bacterially produced K8.1. Heat map analysis of KS patient antibody titers revealed marked heterogeneity in humoral responses to this four-antigen panel. Overall, the LIPS assay showed 97% sensitivity, and positive anti-v-cyclin antibodies were detected in approximately 75% of the KS sera. These results suggest that LIPS screening using an antigen mixture is a sensitive and high-throughput method for serological screening of HHV-8 infection in individuals with KS.

* Corresponding author. Mailing address: Building 49, Room 1C20, 49 Convent Drive, Bethesda, MD 20892-4410. Phone: (301) 402-0778. Fax: (301) 402-0667. E-mail: burbelop{at}nidcr.nih.gov

{triangledown} Published ahead of print on 4 March 2009.

Clinical and Vaccine Immunology, May 2009, p. 621-627, Vol. 16, No. 5
1071-412X/09/$08.00+0     doi:10.1128/CVI.00474-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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