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A fast and simple dot-immunobinding assay for quantifiction of mouse immunoglobulins in hybridoma culture supernatants
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SYSNO ASEP 0105288 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Ostatní články Title A fast and simple dot-immunobinding assay for quantifiction of mouse immunoglobulins in hybridoma culture supernatants Title Rychlý a jednoduchý imunovazebný test ("dot-immunobinding assay") pro kvantifikaci myších imunoglobulinů v supernatantech hybridomových kultur Author(s) Sulimenko, Tetyana (UMG-J)
Dráber, Pavel (UMG-J) RID, ORCIDSource Title Journal of Immunological Methods. - : Elsevier - ISSN 0022-1759
Roč. 289, - (2004), s. 89-95Number of pages 7 s. Language eng - English Country US - United States Keywords dot-immunobinding assay ; hybridoma culture superntatants ; mouse immunoglobulins Subject RIV EB - Genetics ; Molecular Biology R&D Projects IBS5052301 GA AV ČR - Academy of Sciences of the Czech Republic (AV ČR) LN00A026 GA MŠMT - Ministry of Education, Youth and Sports (MEYS) 1P04OE158 GA MŠMT - Ministry of Education, Youth and Sports (MEYS) Annotation Mouse monoclonal antibodies of IgG subclasses and IgM class in hybridoma culture supernatants were quantified using a dot-immunobinding assay. Immunoglobulins were bound to nitrocellulose (NC) membrane and, after blocking, the membrane was incubated with anti-mouse antibody conjugated to horseradish peroxidase (HRP). Binding was revealed by incubation with a sensitive chemiluminiscence reagent. Quantitation was achieved by densitometric comparison with standard curves produced by purified monoclonal antibodies of the same subclass or purified antibodies of the same clone as the antibody to be quantified. These quantitative results were compared with those obtained using purified IgG from mouse serum or purified mouse myeloma IgM as standards. The dot-immunobinding assay requires 1 l of hybridoma culture sample and takes about 1 h in total. Good linearity between the staining intensity and the amount of immobilized immunoglobulins was observed over the range of 0.05ů5 ng/spot. The assay is simple, reproducible and can process simultaneously a large number of samples Workplace Institute of Molecular Genetics Contact Nikol Škňouřilová, nikol.sknourilova@img.cas.cz, Tel.: 241 063 217 Year of Publishing 2005
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