A systematic study of the effect of procedural variables on the ALA-D assay has been carried out. Because of metal content in vacutainers, artifactual results are obtained on blood samples drawn by this method and the method of choice is the use of plastic syringes and tubes. The use of isopropyl alcohol as a skin sterilizing agent, although it gives measurable blood alcohol readings, does not affect the ALA-D assay. The handling of the whole blood has been investigated and the two methods of hemolysis compared. Trichloroacetic acid, a reagent used in the final assay of porphobilinogen, has been found to be a serious interference in the method, making it essential for this reagent to be kept in a securely sealed container. The deterioration of the trichloroacetic acid--mercuric chloride reagent is also a potential procedural problem that must be considered.