Localization of the binding sites of native and acetylated low-density lipoprotein (LDL) in human monocyte-derived macrophages.

Human monocyte-derived macrophages were demonstrated to have separate and morphologically distinct binding sites for low density lipoprotein (LDL) and acetylated LDL (AcLDL). Using an indirect immunoperoxidase technique and electron microscopy, only LDL was shown to bind to its receptor in coated pits on the macrophage membrane, whereas the distribution of AcLDL-receptor complexes was dependent upon whether or not the cells were fixed prior to incubation with AcLDL. In cells incubated with AcLDL, then fixed, electron-dense precipitate was found in aggregates, sometimes near pseudopodia; fixed cells incubated with AcLDL had electron-dense precipitate more uniformly spread along the membrane. These data suggest that the 'scavenger' receptor is diffusely distributed in the membrane and that following AcLDL binding the receptors cluster in regions of the membrane which do not contain coated pits.

Purification of soluble complement-fixing antigens from two Burkitt's lymphoma cell lines. Conditions affecting the stability and recovery of antigens and antibodies.

Soluble Epstein--Barr virus (EBV) associated antigens of P3HR-1 and Raji cell lines, which fix complement in the presence of human sera containing antibody against EBV--VCA antigens, were partially purified by fractionating centrifuged cell lysates by the method of immunoadsorbents or by precipitation with ammonium sulphate or glycine--HCl. Purification conditions were limited by the instability of both antigen and antibody activity. The antibody was inactivated under all the conditions commonly used to dissociate antigen--antibody complexes. The antigen was inactivated at pH 2-3, and was irreversibly bound to Sephadex G-200 conjugates which had not been exposed to large excesses of serum.

Stability and dissociation of P3H4-1 Burkitt's lymphoma cell soluble complement-fixing antigen identified with human serum.

The soluble complement-fixing antigen of the P3HR-1 Burkitt's lymphoma cell line, identified with human serum containing antibody against Epstein-Barr virus viral capsid antigen, loses activity under a variety of conditions. The major characteristic reported here is retention of activity on exposure to specific physical or chemical conditions followed by loss of activity after subsequent neutral dialysis. Antigen of untreated cell lysates, which retains activity after dialysis against large volumes of neutral buffers or 0.14 M NaCl, will lose activity if the lysate is first heated to 56 degrees or if the lysate is exposed to acid perchlorate, and the resulting precipitate and supernatant are redialyzed separately to neutral pH. The P3HR-1 soluble complement-fixing antigen appears to be an aggregate including a labile, dissociable component.

Differences between soluble complement-fixing antigens derived from P3HR-1, RAJI, and Hk-Ly-28 cell lines.

Soluble complement-fixing (CF) antigens of the virus-producing and virus-nonproducing P3HR-1 and RAJI Burkitt lymphoma cell lines and Hk-Ly-28 nasopharyngeal cancer cell line, identified by use of viral capsid antigen-positive human sera, could readily be distinguished by differences in their stability to chemical and physical conditions. The P3HR-1 soluble CF antigen lost titer when heated or exposed to acid perchlorate under conditions in which RAJI and Hk-Ly-28 soluble antigens were stable. Differences in solubility were also found. These results contribute not only to the interpretation of the relationship of Epstein-Barr virus (EBV)-associated components of RAJI to P3HR-1 and Hk-Ly-28 cell lysates but also to the selection of conditions for the development of identification tests and purification procedures for CF antigens and antibodies of Burkitt's lymphoma, nasopharyngeal cancer, and other EBV-associated tumors.