title: Development of a sensitive radioimmunoassay for Fab fragments: application to Fab pharmacokinetics in humans.
abstract: Anti-sheep Fab fragment antisera were produced in rabbits using sheep digoxin-specific Fab fragments (Digidot) as immunogen. These antisera were used for the development of a radioimmunoassay (RIA) of sheep Fab fragments in human plasma and urine using 125I-labeled Fab fragments. Interference in the assays by digoxin, human proteins, and antibodies from different species was insignificant, but cross-reactivity between anti-sheep Fab antisera and goat IgG or Fab fragments was 22 to 67%. The limit of detection was 0.1 microgram/mL and the assay was linear over a 0.6-28 micrograms/mL range of Fab fragments. Intra- and interassay coefficients of variation were less than 6.9 and 10.5%, respectively. Accuracy of plasma and urine assays at various Fab fragment levels ranged from 96 to 106%. RIA was applied to the pharmacokinetic study of sheep digoxin-specific Fab fragments in one patient acutely intoxicated by digitoxin and treated with Digidot. The Fab elimination half-life was 12.1 hr. Steady-state volume of distribution and total-body clearance were 10.8 L and 23.4 mL/min, respectively. Unchanged Fab fragments (50 kD) and degradation products (25 kD) isolated by gel filtration chromatography of a urine sample cross-reacted with the anti-Fab antiserum.
authors.affiliation: INSERM U26, Hôpital F. Widal, Paris, France.
authors.forename: C V
authors.initials: CV
authors.lastname: Thanh-Barthet
authors.forename: M
authors.initials: M
authors.lastname: Urtizberea
authors.forename: A E
authors.initials: AE
authors.lastname: Sabouraud
authors.forename: N J
authors.initials: NJ
authors.lastname: Cano
authors.forename: J M
authors.initials: JM
authors.lastname: Scherrmann
mesh_terms: D000818:Animals; D004074:Digitoxin / Q000506:poisoning; D004077:Digoxin / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006207:Half-Life; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007140:Immunoglobulin Fab Fragments / Q000097:blood* / Q000652:urine*; D008875:Middle Aged; D011863:Radioimmunoassay; D012680:Sensitivity and Specificity
chemical_list: D007106:Immune Sera; D007140:Immunoglobulin Fab Fragments; C050199:digoxin antibodies Fab fragments; D004077:Digoxin; D004074:Digitoxin