Heparin in plasma samples causes nonspecific binding to histones on Western blots.
We observed an artifactual reactivity on Western blots when heparin was used as the anticoagulant in collected blood specimens. This nonspecific interaction was found to be due to immunoglobulin aggregates that bound to cellular proteins, in particular histones. Nonspecific interaction was not observed in fresh heparinized samples, but was present in samples frozen for long-term storage. Other anticoagulants such as EDTA, oxaloacetate and sodium citrate did not cause this nonspecific reactivity. Although adding heparin to serum could reproduce the nonspecific reactivity on Western blots, other immunological tests such as ELISA or indirect immunofluorescence were not affected by the use of heparinized plasma. Enzymatic digestion of heparinized samples with Heparinase I removed the artifactual reactivity, leaving specific antigen-antibody interactions unaffected. Therefore, we advise caution in the interpretation of Western blotting experiments when blood or other tissue fluid specimens are collected in heparin.
Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA.
F F
FF
Alcantara
D J
DJ
Iglehart
R L
RL
Ochs
D000925:Anticoagulants / Q000097:blood / Q000378:metabolism*; D015153:Blotting, Western / Q000379:methods*; D006493:Heparin / Q000097:blood / Q000378:metabolism*; D006657:Histones / Q000378:metabolism*; D006801:Humans; D014407:Tumor Cells, Cultured
D000925:Anticoagulants; D006657:Histones; D006493:Heparin