Development of a microwell adapted immunoblot system with recombinant antigens for distinguishing human herpesvirus (HHV)6A and HHV6B and detection of human cytomegalovirus

Clin Chem Lab Med. 2011 Nov;49(11):1891-8. doi: 10.1515/CCLM.2011.666. Epub 2011 Jul 14.

Abstract

Background: The human cytomegalovirus (HCMV) and the human herpesvirus 6 (HHV6) are widely distributed in the human population. The variants A and B of HHV6 are closely related to each other and cannot be distinguished by common serological methods like enzyme-linked immunosorbent assay (ELISA) or immunofluorescence test (IFT). The aim of this study was to develop a microwell-adapted blot system for specificity detection of human cytomegalovirus and human herpesvirus 6A and 6B (HHV6A, HHV6B) that combines the advantages of ELISA (automation and multiplex detection) and immunoblotting (antigen-specific antibody detection with high specificity).

Methods: Ten HCMV, five HHV6A and five HHV6B antigens were expressed as fusion proteins and tested with sera of children (n=30), of healthy young adults (n=30) and of older adults (n=30) in a newly developed microblot system.

Results: Sensitivity and specificity of HCMV and HHV6 microblots were comparable to commercially available[ELISA, IFT and to line assay tests. The advantage of the HHV6 microblot is the possibility of distinguishing between HHV6A-monovalent sera, HHV6B-monovalent sera and HHV6A/B-polyvalent sera. Most sera of children younger than 2 years showed only HHV6B antigen positivity, while most sera of adults and children aged over 2 years reacted with HHV6A and B proteins, although predominance for HHV6B was observed.

Conclusions: The authors were able to detect HCMV positive sera and to distinguish between HHV6A-monovalent sera, HHV6B-monovalent sera and HHVA/B-polyvalent sera with the new developed microblot system. Predominance of HHV6B was observed in sera of children and adults.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Antibodies, Viral / immunology
  • Antigens, Viral / immunology
  • Automation, Laboratory
  • Child
  • Cloning, Molecular
  • Cytomegalovirus / genetics
  • Cytomegalovirus / immunology
  • Cytomegalovirus / isolation & purification*
  • Cytomegalovirus Infections / diagnosis*
  • Cytomegalovirus Infections / immunology
  • Cytomegalovirus Infections / virology
  • Enzyme-Linked Immunosorbent Assay / methods*
  • Escherichia coli
  • Female
  • Herpesvirus 6, Human / genetics
  • Herpesvirus 6, Human / immunology
  • Herpesvirus 6, Human / isolation & purification*
  • Humans
  • Immunoblotting / methods*
  • Male
  • Plasmids
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Recombinant Proteins / isolation & purification
  • Roseolovirus Infections / diagnosis*
  • Roseolovirus Infections / immunology
  • Roseolovirus Infections / virology
  • Sensitivity and Specificity
  • Transformation, Bacterial
  • Young Adult

Substances

  • Antibodies, Viral
  • Antigens, Viral
  • Recombinant Proteins