Rapid and inexpensive enzyme inhibition assay of methotrexate.

A rapid and inexpensive enzyme inhibition procedure has been developed for the assay of methotrexate in biological fluids. The method achieves a high degree of sensitivity, reproducibility, and specificity. No prior separation procedures are required, and there is no cross-reaction with metabolites of methotrexate or other substances such as folinic acid, which is often given concurrently with methotrexate. Results are in good agreement with those given by the more expensive enzyme multiplied immunoassay technique. The new method is 30 times more sensitive than the enzyme multiplied immunoassay technique, being capable of determining concentrations as low as 1.0 X 10(-8) M (4.5 ng/ml), a distinct advantage over the enzyme multiplied immunoassay technique, since it may be successfully employed in conducting clinical pharmacokinetic studies.