Rapid procedure for fecal porphyrin assay

Hydrochloric acid extraction of feces in the presence of ether yields an extract suitable for spectrophotometric estimation of total porphyrin and for further separation by "high-performance" liquid chromatography (HPLC) or thin-layer chromatography. A total porphyrin reference interval of less than 200 nmol/g dry weight of feces was established from data on 106 normal subjects on an unrestricted diet. Total fecal porphyrin values in 11 porphyria cutanea tarda patients were considerably higher than given by the widely used Rimington method (respective means, 652 and 239 nmol/g dry weight). Our HPLC method for separation of porphyrin methyl esters on a silica column, with quantification by fluorescence, is described. HPLC separations performed on 23 porphyria cutanea tarda patients gave the following mean proportions of total fecal porphyrins: dicarboxylics 21%, coproporphyrin 9%, isocoproporphyrins 28%, pentacarboxylporphyrin 9%, hexacarboxylporphyrin 11%, heptacarboxylporphyrin 18%, and uroporphyrin 4%.     

An improved technique for immunofixation of electrophoretograms.

Immunofixation coupled with electrophoresis is winning favor as an easy, sensitive, and versatile method for detecting specific protein fractions. Here, we discuss problems which appear when antisera are applied onto the electrophoretic medium (Cellogel or cellulose acetate). Longitudinal slits can be made in the electrophoretic medium without worsening resolution. Such longitudinally slitted electrophoretic membranes can be used in electrophoresis and specific antisera applied simply by pipetting them onto the electrophoretograms. The method has been used successfully in typing immunoglobulins.     

Derivative electrophoretograms and their applications.

A differentiator was constructed to produce the first and second derivative curves of routine cellulose acetate membrane electrophoretograms of human serum protein. The derivative curves could reveal the presence of multicomponent in each globulin fraction. The "M" components could be detected easily and semiquantitatively, when the cellulose acetate membrane electrophoretic pattern was evaluated by the derivative curves. Utilization of the derivative curves could be a useful adjunct to the detection of an "M" component since the presence of a narrow band produces a sharper peak on the second derivative curve and more indentation on the first derivative curve.     

Rapid radioisotopic procedure for determination of nortriptyline in plasma.

With the widespread use of tricyclic antidepressant drugs, the relationship between the concentration of the drug in the plasma and the therapeutic response is of considerable interest. We describe a double-isotope derivative dilution procedure for measuring plasma nortriptyline. In the method, [14C]nortriptyline is used for estimating procedural losses and [3H]acetic anhydride for derivative formation. The assay is rapid and adequately specific, sensitive, precies, and reproducible for routine clinical use. We used it to investigate the variation in steady-state drug concentrations in plasma of persons who were on a 150 mg/day dose of nortriptyline. Intra-individual variation from day to day was 10-14%. This variation was not significantly affected by the dosage schedule, the time of sampling after an oral dose, or the storage of the plasma samples. For 19 patients on 150 mg of nortriptyline per day, the mean concentration in plasma was 181 +/- 22 (SE) mug/liter, a value that compares well with our previous findings and those of other groups.     

Procedure for characterization of creatine kinase variants on agarose electrophoretograms

Electrophoretograms of atypical creatine kinase (EC 2.7.3.2, CK) isoenzymes may be difficult to interpret. In addition, their presence may cause discrepancies between results by immunological determinations for mass or activity and those by electrophoresis. Here we outline procedures to use in characterizing apparently atypical CK isoenzymes observed after electrophoresis, as illustrated by a CK isoenzyme variant that migrated as CK-MB.     

A modified automated fluorometric method for tyrosine determination in blood spotted on paper: a mass screening procedure for tyrosinemia

A modified version of the Hochella [2] automated fluorometric technique for tyrosine determination is presented as a mass screening procedure for tyrosinemia. With this technique, 85 000 tests a year can be done with one Auto-Analyzer.In the province of Quebec, 17 cases of hereditary tyrosinemia have been detected with this method over the past three years (208 000 tests).Although the number of false positives is high, blood tyrosine determination remains, to our knowledge, the only mass screening procedure available to detect hereditary tyrosinemia.