Enhanced Clearance of Specifically Bound Digoxin from Human Myocardial and Skeletal Muscle Samples by Specific Digoxin Antibody Fragments

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Abstract
The effect of digoxin antibody fragments (F on clearance of specifically bound digoxin from its specific receptor Na.K-A [Pase) was studied in human heart left ventricle (LV) and vastus lateras skeletal muscle (SK) samples obtained postmortem. Initially. [H] digoxin was hound to samples it conditions giving high relative occupancy of receptors. Half-life it's for its net release from LV in buffer was 32.2. 6.7. and 0.9 hat 0.30 and 37C. respectively. For SK U was 5.4 h in buffer at 30 C. Inhibition of rebinding of digoxin by addition of specific digoxin 15 10 M) or excess unlabeled digoxin (, 15 ± 10 M) or excess unlabeled digoxin (10 M) to buffer at 30 C increased net release rate for specifically bound samples at condition giving low relative occupancy. Samples were suhsequently washed in buffer containing 5 10 M specific digoxin 1 for 16 at 30 C. This wash reduced occupancy of receptors by digoxin from 10 to 0.5% in LV and from 9 to 0.3 in SK, respectively. At variance With wash at 37. C this procedure allowed subsequent vanadate-facilitated complete quantification of Na.KAT-Pase by ['H]ouahain binding: values were 378 ± 13 and 370 ± 12 pmol g wet weight (p 0.6) in LV and 309 ± 19 and 315 ± 16 pmol o wet weight (p 0.7) in SK With and without previous wash. respectively (means - SEM. n 12). [H] Ouabain binding to samples without prior digoxin binding was also unaffected by the wash. Thus. it may he beneficial in treatment of digitalis intoxication to ensure sustained high levels of free F in the blood for a time: no evidence was found to support the existence of an endogenous digitalis-like factor: the developed method may prove to he of value fur accurate determinations of Na.K-ATPase in heart and SK samples from digitalized patients.

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