Comparison study of streptavidin-coated microtitration plates.
Streptavidin (SAv)-coated 96-well microtitration plates from commercial suppliers (six plates) and our laboratory (two plates) were tested with respect to their binding capacity for small and large molecules, leaching, well-to-well variation and immunoassay performance. The binding capacities for europium-labelled biotin (Eu-biotin) varied from 4.4 to above 150 pmol/well depending on the plate, and for biotinylated monoclonal antibody from 1.2 to 6.4 pmol/well (190-1030 ng). Incubation for 1 h in regular immunoassay buffer resulted in leaching of 0.6-76 ng of SAv from unwashed wells and 0.5-60 ng from pre-washed wells. The desorbed quantity represented up to 3.4% of the maximal Eu-biotin binding capacity. Coating-related variation of 96 wells (CV% values) ranged from 1.2% to 8.0% when tested with respect to maximum binding capacity. Immunofluorometric assay for TSH as well as enzyme immunoassays for CA125 and PSA revealed immunoassay-related performance of the plates, regarding signal levels, variation and non-specific binding characteristics. Non-specific binding and variation tended to increase in the highest capacity plates, whereas some low capacity plates fulfilled the assay requirements optimally. Good performance with respect to capacity, coating homogeneity or leaching was not by definition reflected in the performance of two-site heterogeneous immunoassays.
Välimaa
Lasse
L
University of Turku, Department of Biotechnology, Tykistökatu 6 A, FIN-20520 Turku, Finland. lasse.valimaa@utu.fi
Laurikainen
Katja
K
D000911:Antibodies, Monoclonal; D001710:Biotin; D018394:CA-125 Antigen / Q000032:analysis; D005063:Europium; D005260:Female; D015200:Fluoroimmunoassay / Q000295:instrumentation / Q000379:methods; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods*; D007124:Immunoenzyme Techniques / Q000295:instrumentation / Q000379:methods; D008297:Male; D017430:Prostate-Specific Antigen / Q000032:analysis; D019809:Streptavidin; D013972:Thyrotropin / Q000032:analysis
D000911:Antibodies, Monoclonal; D018394:CA-125 Antigen; D005063:Europium; D001710:Biotin; D013972:Thyrotropin; D019809:Streptavidin; D017430:Prostate-Specific Antigen