Dot immunodetection for sphingomyelinase with monoclonal antibody.
An assay for the detection of sphingomyelinase with monoclonal antibodies is described. The assay takes advantage of nitrocellulose membranes as antigen adsorbent on which a dilute sample can be concentrated as a spot, using a specially designed 96-well filtration device which is commercially available. The method requires only 6 micrograms of the extracts from leukocytes and liver, and it is 10 times more sensitive than the colorimetric assay. This reduced amount of sample material also has the merit of requiring only a 0.5-ml blood sample from patients. The combination of this dot immunoassay with the monoclonal antibody allows a sensitive and a specific assay, and is also applicable as a screening test on a large number of samples. Furthermore, the possibility of differential diagnosis of Niemann-Pick disease types by detecting isoenzymes by this method was examined after isoelectric focusing of placental sphingomyelinase.
D000818:Animals; D000911:Antibodies, Monoclonal; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D007118:Immunoassay / Q000379:methods; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000276:immunology / Q000378:metabolism*; D007962:Leukocytes / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D051379:Mice; D008807:Mice, Inbred BALB C; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D010920:Placenta / Q000201:enzymology; D013108:Sphingomyelin Phosphodiesterase / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D007527:Isoenzymes; D010727:Phosphoric Diester Hydrolases; D013108:Sphingomyelin Phosphodiesterase