We have developed a double antibody radioimmunoassay (RIA) for human apolipoprotein B (ApoB). The assay measures not only the ApoB content of beta-lipoproteins (low density lipoproteins [LDL]) but also that contained in the other lipoproteins in plasma. Purified lymph and plasma chylomicrons and plasma very low density lipoproteins (VLDL) produced displacement curves in the assay system which paralleled those produced by pure LDL. Thus, the ApoB found in chylomicrons, VLDL, and LDL were immunologically identical. ApoB accounted for about 25 and 35%, respectively, of the total protein of chylomicrons and VLDL by RIA. VLDL and LDL preparations from normal and hyperlipoproteinemic subjects also produced parallel displacement curves, suggesting that the ApoB of normal and hyperlipoproteinemic subjects were immunologically identical. High density lipoproteins and abetalipoproteinemic plasma displaced no counts, nor did the sera of several animal species produce any useful displacement curves in this system. The fasting total plasma ApoB concentration of normal subjects was 83+/-16 mg/dl (mean+/-SD). ApoB levels were high in Type II (162+/-16), and less so in Type IV (112+/-24) and Type V (105+/-17).When plasma ApoB concentration in Type IV patients was graphed against plasma glycerides, two subpopulations, which may represent different genetic or biochemical subgroups, were apparent.ApoB concentration in individuals on constant diet and drug regimen was stable over weeks to months. Greater than 90% of ApoB of normal and Type II subjects was in the d > 1.006 plasma fraction. By contrast, only 50-80% of ApoB was in the d > 1.006 fraction in Types IV and V. Thus, hypertriglyceridemia was associated primarily with a redistribution of ApoB to the lighter density fractions; by contrast, in hypercholesterolemia absolute ApoB concentration was markedly increased.