ISSN:
1420-9071
Keywords:
Human
;
leucocytes
;
myeloperoxidase-iodination-radioimmunoassay
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Medicine
Notes:
Abstract In order to obtain a radioimmunoassay (RIA) technique for the measurement of human plasma myeloperoxidase (MPO), we purified the enzyme from polymorphonuclear granulocytes (neutrophils), and compared three methods of labeling it with125Iodine: chloramine T, lactoperoxidase, and an original technique of ‘self labeling’ based on the ability of the enzyme to oxidize and bind125I in the presence of H2O2. The chloramine T technique produced a degraded protein, as well shown by a high non-specific binding of tracer to antibody. The lactoperoxidase technique did not succeed in labeling MPO with an adequate specific activity. In contrast, the self-labeling method gave a stable tracer with a specific activity of 23 μCi/gmg MPO (85 MBq), a satisfactory level of immunoreactivity, and a low-specific binding (≤3%). After labeling, purification of tracer was achieved by gel filtration chromatography in phosphate buffer (0.05 M; pH7) to which 0.1% poly-L-lysine was added. The labeled molecule remained stable for 40 days and could be used for RIA with a polyclonal antibody raised in rabbits.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF01929890
Permalink
