Measurement of cytotoxic T lymphocyte activity of human cytomegalovirus seropositive individuals by a highly sensitive coupled luminescent method.
A coupled luminescent method (CLM) based on glyceraldehyde-3-phosphate dehydrogenase released from injured target cells was used to evaluate the cytotoxicity of antigen-specific HLA class I-restricted CTLs. In contrast to established methods, CLM does not require the pretreatment of target cells with radioactive or toxic labeling substances. CTLs from healthy HLA-A2 positive donors were stimulated by autologous dendritic cells (DCs) pulsed with HLA-A2 restricted HCMV-pp65 nonamer peptides. HLA-A2 positive T2 cells or autologous monocytes pulsed with HCMV-pp65 nonamer peptide served as target cells. Lysis was detected only in HCMV-pp65-pulsed target cells incubated with CTLs from seropositive donors stimulated by HCMV-pp65-pulsed DCs. After 3 days, stimulation 38% of T2 cells and 17% of monocytes were lysed at an effector to target ratio of 8:1. In conclusion, CLM represents a highly sensitive, fast, material-saving and non-toxic/non-radioactive method for the measurement of antigen-specific CTL cytotoxic activity.
Ogbomo
Henry
H
Institut für Medizinische Virologie, Zentrum der Hygiene, Klinikum der Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany.
Geiler
Janina
J
Leutz
Anke
A
von Kietzell
Kristina
K
Michaelis
Martin
M
Doerr
Hans Wilhelm
HW
Cinatl
Jindrich
J
D003587:Cytomegalovirus / Q000276:immunology; D003586:Cytomegalovirus Infections / Q000276:immunology*; D003713:Dendritic Cells / Q000276:immunology; D015789:HLA-A2 Antigen / Q000276:immunology; D006801:Humans; D008163:Luminescent Measurements / Q000379:methods*; D008213:Lymphocyte Activation; D010750:Phosphoproteins / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014763:Viral Matrix Proteins / Q000276:immunology
D015789:HLA-A2 Antigen; D010750:Phosphoproteins; D014763:Viral Matrix Proteins; C065418:cytomegalovirus matrix protein 65kDa