Flow immunochemical bio-recognition detection for the determination of interleukin-10 in cell samples.
On- and off-line heterogeneous non-competitive flow immunoassays for the determination of Interleukin-10 are described. The sample containing IL-10 is mixed, either on-line in a reaction coil or off-line in a test tube, with fluorescent labelled anti-IL-10 antibodies to form an antibody-antigen complex. The labelled unbound antibodies are trapped on an immobilized IL-10 column whereas the IL-10-antibody complexes are eluted and detected downstream by a fluorescence detector. The optimization of the systems was performed with respect to choice of affinity support, flow rate, carrier buffer additives, pH and antibody-antigen association. Both bio recognition assays were tested with a spiked cell medium and the IL-10 detection limits in this matrix was found to be 8 fmol using the off-line incubation mode and 40 fmol using the on-line incubation mode. The sample through-put was 26 and 40 samples per hour in the on-line and off-line incubation modes, respectively. IL-10 identification in the sample fractions was achieved using MALDI-TOF MS.
Kjellström
S
S
Department of Analytical Chemistry, University of Lund, P.O. Box 124, S-221 00, Lund, Sweden.
Emnéus
J
J
Marko-Varga
G
G
D000937:Antigen-Antibody Reactions; D002846:Chromatography, Affinity; D003429:Cross Reactions; D015200:Fluoroimmunoassay / Q000379:methods*; D006801:Humans; D016753:Interleukin-10 / Q000032:analysis*; D007700:Kinetics; D011994:Recombinant Proteins / Q000032:analysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
D011994:Recombinant Proteins; D016753:Interleukin-10