Expression of tegument protein pp65 of human cytomegalovirus (CMV) and its application to the analysis of viral-specific cellular immunity in CMV-infected individuals.
Investigations into human cytomegalovirus (CMV)-specific cellular immunity are important to better understand and manage CMV infections. CMV phosphoprotein pp65 is thought to be a major antigen for CMV-specific cellular immunity. We newly synthesized protein pp65 with a baculovirus expression system and purified it via metal affinity chromatography in a soluble form. The recombinant protein pp65 was antigenic in an enzyme immuno-linked assay for pp65-specific IgG in sera from 196 children. Traditional lymphoproliferation assays have shown that pp65 protein promotes specific lymphoproliferation in CMV-seropositive donors. Using an intracellular cytokine detection system, we showed that this recombinant protein stimulated CD4-positive T cells to express interferon-gamma. The results of these assays using protein pp65 were comparable with the use of CMV whole antigen. pp65- and CMV-specific cellular immunity, and CMV DNA load were also compared in four recipients of unrelated cord blood transplantation. The delay in re-constitution in CMV-specific cellular immunity was associated with reactivation of CMV. These results indicated that the recombinant protein pp65 can be used to study specific immunity in CMV infections.
Tanaka
N
N
Department of Pediatrics, Developmental Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan. ntanaka@med.nagoya-u.ac.jp
Kimura
H
H
Hoshino
Y
Y
Nishikawa
K
K
Kojima
S
S
Nishiyama
Y
Y
Morishima
T
T
D000293:Adolescent; D000956:Antigens, Viral / Q000096:biosynthesis / Q000276:immunology; D016367:Baculoviridae / Q000235:genetics; D016913:Blood Component Transfusion; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D003587:Cytomegalovirus / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003586:Cytomegalovirus Infections / Q000097:blood / Q000276:immunology* / Q000628:therapy; D004279:DNA, Viral / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005822:Genetic Vectors; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007223:Infant; D007371:Interferon-gamma / Q000032:analysis; D008297:Male; D010750:Phosphoproteins / Q000096:biosynthesis* / Q000276:immunology; D011994:Recombinant Proteins / Q000096:biosynthesis; D019562:Viral Load; D014763:Viral Matrix Proteins / Q000096:biosynthesis* / Q000276:immunology
D000956:Antigens, Viral; D004279:DNA, Viral; D007074:Immunoglobulin G; D010750:Phosphoproteins; D011994:Recombinant Proteins; D014763:Viral Matrix Proteins; C065418:cytomegalovirus matrix protein 65kDa; D007371:Interferon-gamma