Abstract. Sera from 11 individuals which were classified as Lp(a-) by direct gel diffusion and by absorption tests showed the presence of Lp(a) lipoprotein when the sera were concentrated 120-fold. This Lp(a) antigen was demonstrated by gel diffusion to be indistinguishable from Lp-lipoprotein isolated from known Lp(a+) sera. The Lp(a) preparations from sera classified Lp(a-) also showed electrophoretic mobility in immunoelectrophoresis and schlieren diagrams in ultracentrifugal analyses similar to Lp-lipoprotein from Lp(a+) sera. The proposal is made that observed individual variations in tests for the Lplipoprotein reflect a quantitative genetic trait and that it is likely that different individuals produce Lp-lipoprotein in widely varying amounts. The consistency of this proposal with certain previous observations on the Lp system is discussed.

Small angle x-ray diffraction patterns have been obtained from ordered arrays of hemoglobinfree human erythrocyte membranes by use of improved techniques. Diffraction data have been recorded to 9 A resolution on samples whose lattice periodicity was varied (by changing humidity) from 55.5 A to 69.6 A. The observed reflections permitted tracing the intensity trans. Small angle x-ray diffraction techniques have been used to investigate the erythrocyte membrane (1-7). The diffraction data that have been reported consist of three or four orders of '110-A periodicity for erythrocyte membranes that contain large amounts of hemoglobin and two orders of '70-A periodicity for hemoglobin-free erythrocyte membranes. These data are insufficient to determine phase angles of the reflections by swelling techniques (8), and thus do not permit computation of projected electron density distributions by Fourier inversion. We now report higher resolution x-ray diffraction data from hemoglobin-free human erythrocyte membranes and the computation of projected electron density distributions based upon an assignment of phase angles. MATERIALS AND METHODSWhole human blood, type 0, Rh positive, was drawn into Fenwal JH-IN heparinized blood packs. Hemoglobin-free membranes were prepared by the procedure of Dodge et al. (9). Membranes were hemolyzed and washed with pH 7.4, 15 imOsm sodium phosphate buffer. The membranes were finally suspended in pH 6.3, 15 imOsm phosphate buffer and preserved by the addition of 0.5 mM NaN3. The suspension was examined optically with a Leitz phase contrast microscope and showed a homogeneous preparation of membrane vesicles approximately 6 ,um in diameter.The amount of membrane material in the samples was determined using a modification of the dry weight method described by Hunter (10). A semi-microbalance, Mettler type H16, was used. Membrane samples were measured with class A volumetric pipettes followed by a rinse of the pipette with glass-distilled water into the weighing bottle. The temperature of the vacuum oven was 700 for the second 24-hr period. Dry weights, done in triplicate, were calculated on a weight per volume basis.Total protein was measured by the method of Lowry et al. (11), with bovine serum albumin as a standard. Hemoglobin was determined by a pyridine hemochromagen method (12), with human hemoglobin (Sigma Chemical Co.) as a standard. N-Acetylneuraminic acid (NeuNAc) was measured by the method of Warren (13), with purified NeuNAc (Sigma Chemical Co.) as a standard.Membrane samples were prepared for x-ray diffraction by centrifugation at 105 X g for 24 hr in a Spinco model L ultracentrifuge using a SW 39 rotor at 40. The pellets were equilibrated in a nitrogen atmosphere at 370 and 93-98% relative humidity. The pellets were cut into strips, mounted in the x-ray diffraction chamber, and equilibrated in a water vapor atmosphere at 40 and 75-100% relative humidity before exposure.Diffraction patterns were recorded with a Rigaku Denki RU-3H rotating anode generator, a mirror-monochr...

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Normal human serum and a fraction rich in lipoprotein, Cohn fraction IV,, have been shown in previous studies to detoxify native endotoxin by decreasing lethality for mice, fever in rabbits, and by the alteration of the characteristic endotoxin-anti-endotoxin precipitin pattern in gels. These studies are extended herein and document the ability of normal human serum and fraction IV1 to neutralize the induction of hypotension in rabbits by viable gram-negative bacilli. Further fractionation of serum, using an ultracentrifugal flotation method for producing lipoprotein-free human serum and purified high-density lipoproteins, revealed the lipoprotein-free fraction to be capable of inhibiting endotoxin hypotensive activity and to alter diffusion of endotoxin in gels. On the other hand, the purified high-density lipoproteins failed to negate either activity.

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