Development and application of a radioimmunoassay for methotrexate

Abstract

Summary.-An antiserum to methotrexate has been produced in a sheep against a conjugate of ovalbumin and methotrexate (MTX) prepared using a water-soluble carbodiimide. The antibodies produced were specific for substances containing the 2,4-diamino pteridine structure. Naturally occurring folates did not interfere with the assay. A radioimmunoassay has been developed using this antiserum, which can be used to measure MTX concentrations of less than 1 ng/ml in biological samples without prior extraction. The concentrations of MTX in the blood and urine of patients following a single i.v. bolus injection and following oral administration of the drug have been measured. The published radioimmunoassays for MTX have been compared.

Comparison of plasma and tissue levels of ZD1694 (Tomudex), a highly polyglutamatable quinazoline thymidylate synthase inhibitor, in preclinical models

“…As there is as yet no evidence of plasma and urinary metabolites of ZD1694 (Jodrell et al, 1991;Judson et al, 1996), plasma drug measurements made with the antiserum can be considered to be specific for ZD1694. The remarkable ability of antisera to distinguish between antifolates and natural folates has been shown before (Levine and Powers, 1974;Aheme et al, 1977) and the low cross-reaction of the ZD1694 antiserum with reduced folates was therefore not surprising. From previous work, it is also to be expected (Anzai et al, 1987) that polyglutamated forms of antifolates will be recognized to a similar extent as the monoglutamate.…”

Radioimmunoassay of Methotrexate: Use of 75Se-Labelled Methotrexate

“…The crossreaction of 7-hydroxy-MTX in the case of 75Se-MTX was thought to be due entirely to 2 %contamination by unchanged MTX of this batch of7-hydroxy-MTX which was different from that used with 3H-MTX. Ten micrograms of several chemotherapeutic agents likely to be prescribed with MTX did not affect the binding of 3H-MTX to antibody (Aherne et al, 1977) and it is unlikely that they would crossreact when 75Se-MTX is used. between the methods over an extremely large range of concentrations was obtained.…”

“…The preparation of specific MTX antiserum has been described (Aherne et al, 1977) and the same antiserum bleed HPjSj3 llA has been used for this work. The antiserum was used in the RIA at a final dilution of 1 :24 000.…”

“…Several radioimmunoassays (RIAs) for the measurement of methotrexate (MTX), all of which use tritiated drug as tracer, have been described (Bohuon et al, 1974; Levine and Powers, 1974; Raso and Schreiber, 1975;Hendel et al, 1976;Loeffler et al, 1976; Aherne et al, 1977). RIA techniques for the measurement of MTX are extremely sensitive and are being used increasingly in the clinical situation.…”

A radioimmunoassay for cytosine arabinoside

“…Gas-liquid chromatography and mass spectroscopy have also been used (Boutagy & Harvey, 1978). Immunoassay is probably the best method for routine monitoring for cytotoxic drug levels (Teale et al, 1977;Aherne et al, 1977).…”