Binding of radiographic contrast media to serum proteins demonstrated by immunoelectrophoresis and radioautography.

Abstract

Many attempts have been made to explain the adverse reactions or toxicity of iodinated radiographic contrast media, but the exact mechanisms have yet to be established (6, 12). Recent investigations, however, have suggested an important relationship between serum protein-binding sites and drug reactions (2, 7, 11). Lasser et al. (11) evaluated the binding characteristics of radiographic contrast media by equilibrium dialysis. They showed that these media bind almost entirely to albumin, and the strength of the protein binding correlated with the LD 50 toxicity. However, this method does not allow easy evaluation of protein-binding sites in individual patients. Kutt et al. (8, 9) investigated the effect of iodinized contrast media on the paper electrophoretic mobilities of blood proteins and lipoproteins. They demonstrated marked alterations from normal patterns. We were unable to confirm Kutt's results (even when his technics were used), but we did devise an in vitro method for the evaluation of serum or plasma protein-binding in individual patients. This was undertaken by standard immunologic electrophoretic technics and radioautography. This method was then used to determine if abnormal binding sites were indeed important factors in the production of reactions and if a predictive test for toxicity to iodinated radiographic contrast media could be developed. Method Serum, plasma, whole blood, and saline were combined with contrast media made radioactive with I131 in a concentration of 0.10–0.18 mg per 100 ml and incubated at 37° for one hour. Protein electrophoresis on paper and cellulose acetate was performed by standard methods (5); gel immunoelectrophoresis was carried out by a modification of the method of Scheidegger (15). Twenty-four hours after the addition of antiserum the gel slides were dried at 37°. Radioautographs were made with Type M Kodak film with a ten-day exposure for 50μc of I131 on the slide. Following radioautography, the slides were washed in saline and water and stained with Ponceau S. The radioautographs were then compared to the original slides (Fig. 4). The protein-binding sites were evaluated in a semiquantitative manner. Results It was initially established that there was no alteration in either the paper, the cellulose polyacetate, or the immunoelectrophoretic protein patterns when radiographic contrast media were added to either the serum, whole blood, or plasma. Figure 1 shows normal distribution in a normal patient and the identification of the proteins found in serum. It illustrates many of the protein fractions that can be identified by this method. Figure 2 shows that this distribution was not altered by the addition of Cholografin.