Neuroendocrine differentiation antigen on human lung carcinoma and Kulchitski cells.

In the normal lung, a subset of cells with a histological appearance consistent with that of Kulchitski cells are the only lung cells reacting with a monoclonal antibody (MOC-1) raised against a human small cell lung carcinoma-derived cell line. Outside the lung, a subset of normal endocrine cells (in the adrenal, thyroid, ovary, and pancreas) as well as neural cells (brain and peripheral Schwann cells) also express the antigen detected by MOC-1 (named MOC-1-related antigen). Some of these positively reacting cells are ectodermally derived, whereas others are of proven endodermal origin, indicating that the MOC-1-related antigen is not a cell lineage-specific antigen. Instead, the common expression of the antigen by cells with a neural, endocrine, or neuroendocrine function suggests that the antigen related to a neuroendocrine differentiation state of these cells. The presence of the MOC-1-related antigen on several non-lung tumors mostly paralleled its normal tissue distribution, indicating that the antigen is generally retained upon malignant transformation. In lung carcinoma, the antigen proves to be present on almost all small cell carcinomas tested. In addition, adenocarcinoma and mixed adenosquamous carcinoma could also express the antigen, whereas pure squamous cell carcinoma generally did not. This finding will be discussed in relation to a proposed "common stem cell" histogenesis of lung carcinoma.

Monoclonal Antibodies to the Cell-Wall-Associated Proteinase of Lactococcus lactis subsp. cremoris Wg2.

Twelve monoclonal antibodies directed to the cell-wall-associated proteinase of Lactococcus lactis subsp. cremoris Wg2 were isolated after immunization of BALB/c mice with a partially purified preparation of the proteinase. The monoclonal antibodies reacted with the 126-kilodalton proteinase band in a Western immunoblot. All but one of the monoclonal antibodies reacted with protein bands with a molecular weight below 126,000, possibly degradation products of the proteinase. The monoclonal antibodies could be divided into six groups according to their different reactions with the proteinase degradation products in the Western blot. Different groups of monoclonal antibodies reacted with different components of the L. lactis subsp. cremoris Wg2 proteinase. Crossed immunoelectrophoresis showed that monoclonal antibody groups I, II, and III react with proteinase component A and that groups IV, V, and VI react with proteinase component B. The isolated monoclonal antibodies cross-reacted with the proteinases of other L. lactis subspecies. Monoclonal antibodies of group IV cross-reacted with proteinase component C of other L. lactis subsp. cremoris strains. The molecular weight of the proteinase attached to the cells of L. lactis subsp. cremoris Wg2 was 200,000, which is different from the previously reported values. This could be analyzed by immunodetection of the proteinase on a Western blot. This value corresponds to the molecular weight calculated from the amino acid sequence of the cloned L. lactis subsp. cremoris Wg2 proteinase gene.

Role of environmental antigen in the development of IgG+ cells in the bursa of fabricius.

IgG+ cells were detected in the bursa of Fabricius after hatching by immunofluorescence staining of single cells and immunohistologic studies using mAb anti-Ig gamma-heavy chain. The frequency of IgG+ cells in the bursa increased rapidly immediately after hatching. In histologic studies, most of the IgG+ cells were found in the medullary areas of the bursal follicle. Isolation of the bursa from the gut by bursal duct ligation before hatching suppressed the development of IgG+ cells in the bursa after hatching. In addition, administration of Ags into the bursal lumen just before bursal duct ligation caused a significant increase of IgG+ cells in the bursa compared with the isolated bursa. However, parenteral administration of Ags had no effect on the frequency of IgG+ cells in the isolated and normal bursa. These results suggest that Ags in the bursa are a prerequisite for the development of IgG+ cells in the bursa. Although the majority of IgG+ bursal cells were IgM+ IgG+ double-positive cells, sorted Bu1+ bursal cells, which contained 99.9% of IgG+ cells but not plasma cells, synthesized de novo IgM but little or no IgG. Therefore, it is likely that surface IgG on the bursal cells is not produced locally, but is trapped by IgM+ bursal cells. We speculate that Ags derived from the bursal lumen form immune complexes within the bursa, which are subsequently trapped on the surface of IgM+ bursal cells mediated either by Fc-receptors, C3 receptors, or surface Ig.

Limited proteolysis of the 94 000-dalton subunit of Panulirus interruptus hemocyanin; the carbohydrate attachment site.

Limited proteolysis of the native monomeric 94-kDa subunit of Panulirus interruptus hemocyanin by trypsin, plasmin and subtilisin produces an 18-kDa fragment, a 71-kDa fragment and a small glycopeptide. In the plasmin digest a 23-kDa precursor of the 18-kDa fragment has been observed. Automatic Edman degradations demonstrated that the 18-kDa fragments have their origin at the N terminus of the 94-kDa subunit and incubations with carboxypeptidase A showed that the 71-kDa fragments originate from the C terminus. The glycopeptide is situated in between. The amino acid sequence of the glycopeptide has been determined. Its carbohydrate content accounts for the total carbohydrate of the 94-kDa subunit. All three proteases cleave the subunit at two positions within a very restricted area of the polypeptide chain, which indicates the presence of an exposed loop, carrying the carbohydrate chain, in the corresponding part the native molecule.

Immunoperoxidase staining on frozen tissue sections as a first screening assay in the preparation of monoclonal antibodies directed against small cell carcinoma of the lung.

To prepare monoclonal antibodies against small cell carcinoma of the lung (SCCL), we have used an SCCL-derived cell line as immunogen. A first screening of hybridoma supernatants was performed on frozen tissue sections of a biopsy with histologically proven SCCL involvement. Screening on tissue sections is a valuable technique, especially for the isolation of monoclonal antibodies directed against tumor antigens. A drawback of this procedure is that it is laborious. To circumvent this, we have reduced the number of supernatants to be screened by increasing the number of seeded hybridomas per well. Although this resulted in the growth of 5-20 hybridomas per well, among which there was always at least one that secreted antibodies against 'common antigens', clones that secreted specific antibodies could still be revealed by testing supernatants which were preabsorbed with thrombocytes. This has resulted in the isolation of 7 monoclonal antibodies directed against SCCL associated antigens.

B lymphocyte differentiation in the rat: production and characterization of monoclonal antibodies to B lineage-associated antigens.

Three mouse monoclonal antibodies (mAb) directed against rat B lineage antigens were produced. The mAb, designated HIS14 (IgG1), HIS22 (IgM) and HIS24 (IgG2b), were characterized for binding to lymphoid and nonlymphoid tissues by immunoperoxidase staining of frozen sections and by (double-) immunofluorescence staining of single cell suspensions from lymphoid organs. HIS14 recognized a pan B cell determinant: it reacted with virtually all cells of each anatomic B cell compartment and with about 95% of surface (s)Ig+ cells in thoracic duct lymph and in suspensions of spleen and lymph nodes. HIS22 and HIS24 detected B lineage-associated antigens expressed by major subpopulations of B cells. HIS22 predominantly stained the lymphocyte corona, but not (or weakly) the germinal centers and splenic marginal zones, whereas HIS24 reacted with both corona and germinal center and not (or weakly) with marginal zone. In accordance with this, substantial proportions of sIg+ cells in spleen cell suspensions did not express HIS22 or HIS24 determinants (20% and 27%, respectively). In bone marrow the vast majority of cytomplasmic mu+ pre-B cells were HIS14+ and HIS24+, and up to one third also HIS22+, indicating an appearance of the determinants early in B lymphocytopoiesis. The antigens recognized by HIS14, HIS22 and HIS24 are lost during the final stage of B cell differentiation: none of the mAb bound to plasma cells. As far as detectable, neither cells of myeloid and erythroid lineages in bone marrow nor thymocytes were stained by HIS14, HIS22, or HIS24. In suspensions of peripheral lymphoid organs (spleen and lymph nodes) but not in thoracic duct lymph, HIS14 and HIS24 labeled a small proportion (12% and 14%, respectively) of Ig- cells. HIS22 did not bind to Ig- peripheral lymphocytes. Reactivity of HIS14, HIS22 and HIS24 with nonlymphoid tissues was virtually absent; HIS22 stained the high endothelial venules in lymph nodes and Peyer's patches. As determined by immunoblotting, the antigenic determinants on lymph node cells recognized by HIS14, HIS22 and HIS24 were present on molecules with an apparent molecular mass of 205 kDa, 210 (and 175) kDa and 205 kDa, respectively, which is similar to the molecular mass of the B cell form of the rat leukocyte common antigen. In addition, the antigens recognized by HIS14, HIS22 and HIS24 co-capped with the leukocyte common antigen. This suggests that each of the three mAb recognize determinants present on the B cell form of the leukocyte common antigen.