Efficient hybridoma screening technique using capture antibody based microarrays.
The hybridoma screening is a key step for the successful generation of high-affinity analyte-specific monoclonal antibodies (MAbs), particularly if the target of the antibody is a low-molecular weight analyte. This work presents an advanced screening method that makes use of antibody microarrays generated by contact printing of the hybridoma cell supernatant samples on glass chips initially coated with capture antibodies. The noncompetitive immunoassay is based on the specific binding of an analyte-horseradish peroxidase conjugate and is performed in an automated fashion using a chemiluminescence readout system. Compared to the standard ELISA screening, the work load is reduced due to a higher degree of automation. The quality of the generated data is comparable to data generated by a previously optimized microplate-based immunoassay method. Among a reference set of 373 hybridoma cell supernatant samples, three out of four high-affinity MAbs were identified as true positive, whereas none of the samples were detected as a false positive.
Rieger
Martin
M
Institute of Hydrochemistry, Technische Universität München, Marchioninistrasse 17, 81377 München, Germany.
Cervino
Christian
C
Sauceda
Jimena C
JC
Niessner
Reinhard
R
Knopp
Dietmar
D
D000818:Animals; D000906:Antibodies / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D006735:Horseradish Peroxidase / Q000378:metabolism; D006825:Hybridomas; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods*; D008163:Luminescent Measurements; D051379:Mice; D046228:Microarray Analysis
D000906:Antibodies; D000911:Antibodies, Monoclonal; D006735:Horseradish Peroxidase