A fast and efficient method for quantification of monoclonal antibodies in an ELISA using a novel incubation system.
We have developed a sensitive, reliable, optimized ELISA to measure human IgM monoclonal antibodies using a novel shaking incubator system with short incubation periods of 15 min at 37 degrees C for all stages. The shaking incubator is compared with a static incubator over a range of incubation times and temperatures. For each stage using static incubation conditions the system does not reach a saturation level and the results are inconsistent, unlike the shaking incubator. No 'edge effects' are observed in the shaking system due to even heating from beneath and across the plate. The orbital shaking ensures optimal mixing of reagents which eliminates a diffusion limited reaction rate caused by a depletion of reactants at the solid phase as observed in the static system. The optimized shaking system permits economical use of reagents since the coating antibody can be used at high dilutions.
International Blood Group Reference Laboratory, Southmead Hospital, Bristol, U.K.
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Mushens
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ML
Scott
D000911:Antibodies, Monoclonal / Q000032:analysis*; D004306:Dose-Response Relationship, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007124:Immunoenzyme Techniques; D007075:Immunoglobulin M / Q000032:analysis*; D013997:Time Factors
D000911:Antibodies, Monoclonal; D007075:Immunoglobulin M