Immunometric assay for lutropin (hLH) based on the use of universal reagents for enzymatic labelling and magnetic separation and monitored by enhanced chemiluminescence.
A solid-phase immunometric assay of human lutropin (hLH) is described. Two different anti-hLH antibodies were utilized as capture antibodies, and anti-IgG antibodies covalently coupled to magnetic particles and horseradish peroxidase, respectively, served as 'universal' detection reagents. An anti-hLH antibody raised in rabbits was incubated with a goat anti-rabbit IgG covalently bound to magnetic particles. The resulting complex was added to a separately incubated mixture of hLH and monoclonal anti-hLH antibody. Following incubation, the immunocomplex was sedimented in a magnetic field and the supernatant discarded. Finally a sheep anti-mouse antibody (F(ab')2 fragment) conjugated to horseradish peroxidase as label was added. Following a further incubation, the particles were sedimented in the magnetic field and washed. The hLH content of the sample was quantitated by measuring 'enhanced chemiluminescence'. The sensitivity of the assay was 2.5 +/- 0.9 IU/l (mean +/- SD), the within-run variation ranged from 7.9 to 11%, the between-run variation from 12.9 to 19.8%. Cross-reaction with hFSH or hTSH could not be detected, but was approximately 0.1% with hCG. The results correlated well with those obtained by radioimmunoassay (r = 0.84).
Pazzagli
M
M
Reproductive Endocrinology Research Unit, Karolinska Hospital, Stockholm, Sweden.
Kohen
F
F
Sufi
S
S
Masironi
B
B
Cekan
S Z
SZ
D000888:Antibodies, Anti-Idiotypic; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D006735:Horseradish Peroxidase; D006801:Humans; D007118:Immunoassay / Q000379:methods / Q000592:standards; D007074:Immunoglobulin G / Q000276:immunology*; D007202:Indicators and Reagents; D007700:Kinetics; D008163:Luminescent Measurements; D007986:Luteinizing Hormone / Q000032:analysis* / Q000276:immunology / Q000592:standards; D008280:Magnetics; D010544:Peroxidases; D012015:Reference Standards
D000888:Antibodies, Anti-Idiotypic; D007074:Immunoglobulin G; D007202:Indicators and Reagents; C048844:anti-IgG; D007986:Luteinizing Hormone; D006735:Horseradish Peroxidase; D010544:Peroxidases