An ELISA for the measurement of human leukemia inhibitory factor in biological fluids and culture supernatants.
A new monoclonal antibody-based ELISA for leukaemia inhibitory factor/human interleukin for DA cells (LIF/HILDA) measurements is described. The sensitivity (56 pg/ml after 4 h incubation, 14 pg/ml after 24 h incubation), precision (intra-assays < 5%), reproducibility (interassay < 10%), and accuracy (recoveries, ranging between 98 and 119%, in several fluids) of the assay, plus its excellent performance in dilution tests, and the lack of interference when in the presence of possible cross-reactive substances guarantee accurate cytokine measurement in biological fluids such as serum, plasma, synovial fluid, follicular fluid, urine and culture supernatants. Using the assay, LIF/HILDA was measurable in supernatants after in vitro whole blood stimulation with phytohemagglutinin (PHA), OKT3, and phorbol myristate acetate (PMA) but not with lipopolysaccharide (LPS) or Ca ionophore. LIF/HILDA production was not measurable until after 24 h of culture, when cytokine levels were seen to increase linearly in the supernatant to reach values of up to 40 ng/ml after 96 h of culture. Finally, a good correlation was found (r = 0.96; p < 0.0001; y = 23.1x + 233) between the LIF/HILDA values obtained using the ELISA and DA-1a bioassay.
De Groote
D
D
Department of Endocrinology, University of Liège, Sart Tilman, Belgium.
Fauchet
F
F
Jadoul
M
M
Dehart
I
I
Raher
S
S
Gevaert
Y
Y
Lopez
M
M
Gathy
R
R
Franssen
J D
JD
Radoux
D
D
D000328:Adult; D000911:Antibodies, Monoclonal; D001681:Biological Assay; D001774:Blood Chemical Analysis / Q000379:methods; D003429:Cross Reactions; D016207:Cytokines / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006131:Growth Inhibitors / Q000032:analysis* / Q000096:biosynthesis; D006801:Humans; D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; D008222:Lymphokines / Q000032:analysis* / Q000096:biosynthesis; D008297:Male; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014556:Urine / Q000737:chemistry
D000911:Antibodies, Monoclonal; D016207:Cytokines; D006131:Growth Inhibitors; D015850:Interleukin-6; C507837:LIF protein, human; D053605:Leukemia Inhibitory Factor; D008222:Lymphokines