Monoclonal antibodies against human granulocytes and myeloid differentiation antigens

Monoclonal antibodies (MCA) were obtained by immunizing BALB/c mice with 99% pure granulocytes from normal donors or with a whole leukocyte suspension obtained from a chronic myelogenous leukemia (CML) patient, and then fusing the mouse spleen cells with a 315–43 myeloma cell clone. Four MCA were selected and studied using ELISA, immunofluorescence, cytotoxicity assays, and FACS analysis. Antibodies 80H.1. 80H.3. and 80H.5 (from normals) and 81H.1 (from CML) detected antigens expressed on neutrophils. Antibodies 80H.1 and 80H.3 (lgG) also reacted with monocytes but not with other blood cell subsets. Antibodies 80H.5 and 81H.1 (lgM) were cytotoxic and reacted strongly with most of the cells of the neutrophil maturation sequence. i.e., myeloblasts, promyelocytes, myelocytes, and mature granulocytes. Antibodies 80H.5 and 81H.1 also inhibited BFU-GM and CFU-E. Antigens recognized by 80H.3. 80H.5, and 81H.1 were expressed both on a proportion of cells from HL.60, KG.1, ML.1, and K562 myeloid cell lines, and on a proportion of blast cells isolated from patients with acute myelogenous leukemia. They were not found on lymphoid cell lines or lymphoid leukemia cells. These MCA recognize either late differentiation antigens expressed on mature neutrophils and monocytes (80H.1 and 80H.3) or early differentiation antigens (80H.5 and 81H.1) specific to the granulocytic lineage. They may be useful for a better definition of those antigens specific to hematopoietic stem cells and their relationship with normal or neoplastic hematopoiesis.     

The major histocompatibility complex (CyLA) of the cynomolgus monkey. I. Serologic definition of 21 specificities

Thirty-seven lymphocytotoxic antisera, 27 of which were raised by immunization with skin grafts and blood from partially matched donors, were tested against cells obtained from 218 unrelated animals and 205 offspring from a colony of cynomolgus monkey (Macaca fascicularis). Evidence was obtained for the presence of at least 21 specificities defined by cluster analysis and segregation within families. Allelic relationships between 16 specificities was suggested by segregation patterns, the absence of triplets and statistical analysis of association in the unrelated population sample. The data support a two-locus model, with tentative assignment of seven specificities to the A locus and six to the B locus. That these lymphocyte alloantigens constitute the major histocompatibility complex (MHC) of the cynomolgus monkey is suggested by analogy with other known MHCs and by the increased survival times of skin grafts between paternally matched half sibs compared to haplodistinct full sibs.     

On the association between amyloid fibrils and glycosaminoglycans; possible interactive role of Ca2+ and amyloid P-component.

We have previously reported the specific association of glycosaminoglycans (GAG) and proteoglycans (PG) with amyloid fibrils and characterized the polysaccharides directly extracted from amyloid-laden tissues. In the present study we further elucidate the association between purified amyloid fibrils and GAG/PG with special reference to those GAG/PG associated with amyloid P-component (AP) and the interactive role of Ca2+ ions. Amyloid fibrils were isolated from human hepatic AA amyloid employing water extraction with and without preceding removal of AP, an extrafibrillar protein component of all amyloids, using sodium citrate. GAG/PG co-isolated with the amyloid extracts, with and without AP, were isolated and characterized. Agarose-affinity chromatography of extracts containing AP was performed, and the GAG associated with this extrafibrillary protein were characterized as well. Several different GAG/PG populations were demonstrated in the various extracts. The abolition of calcium-dependent binding markedly influenced the amount of GAG/PG recovered in the fibril extracts, as well as the total amount of amyloid material obtained. Thus, it seems that calcium plays an important role in the association between the fibrils and the sugar moieties, and that a significant fraction of the GAG found in amyloid exhibits a Ca(2+)-dependent fibril-GAG interaction. No significant difference in the proportion between galactosaminoglycans and glucosamines was, however, disclosed when the two extraction protocols were compared, suggesting that no particular GAG species has a higher affinity for the fibrils themselves. Both dermatan/chondroitin sulphate and heparan sulphate identified in the present study exhibited a Ca(2+)-dependent interaction with AP, supporting previous findings. However, the amyloid-associated galactosaminoglycans found, especially the large PG appearing in small amounts, seemed to have a higher affinity for the extrafibrillar AP than the other GAG.     

Genetic polymorphisms of orosomucoid and alpha-2-HS-glycoprotein in Thai,Sri Lankan and Paraguayan populations

The genetic polymorphism of orosomucoid (ORM) and alpha-2-HS-glycoprotein (AHSG) were studied in Thai, Sri Lankan and Paraguayan populations using isoelectric focusing. Gene frequencies in these populations were compared with those in other populations. Four new ORM variants were detected:ORM1 ^* 15 in Thai,ORM1 ^* 16 in Paraguayan,ORM2 ^* 21 andORM2 ^* 22 in Sri Lankan.     

Comparison of radioimmunoassay and ELISA methods for detection of antibodies to chromatin components

A solid phase radioimmunoassay has been compared with an enzyme-linked immunosorbent assay (ELISA) for efficacy in measuring anti-chromatin antibodies. The low backgrounds achieved with the radioimmunoassay method produced a high signal-to-noise ratio and enabled detection of the human test antiserum at a dilution of 1:102,400. By contrast, the ELISA could detect the same antiserum only at a dilution of 1:3200 and above. The radioimmunoassay was consistently more sensitive than the ELISA for detection of anti-chromatin antibodies in a number of human and mouse sera and ascites fluid containing a monoclonal antibody. Factors affecting sensitivity in both assays are discussed.     

An immunodominant haptenic epitope of carbamazepine detected in serum from patients given long-term treatment with carbamazepine without allergic reaction

An anticarbamazepine antibody was detected in the serum of a patient with severe carbamazepine-induced serum sickness. We found that the patient's T cells and IgG antibody recognized an epitope which appeared in subjects showing an allergic reaction, as well as that in subjects who showed no allergic reaction, after long-term carbamazepine therapy. These results show that an anti-carbamazepine immune response does not occur in the majority of subjects who undergo long-term carbamazepine therapy without developing allergic symptoms, although the immunodominant haptenic epitope of carbamazepine is present in their sera.     

胃动素对血管灌流大鼠离体胃运动的作用

通过血管灌流大鼠离体胃,探讨胃动素对胃运动的影响。结果表明:(1)胃动素可以明显地兴奋胃窦自发的胃运动;(2)胃动素抗血清可以完全消除胃动素兴奋胃窦运动的作用;(3)阿托品可以阻断胃动素兴奋胃窦运动的作用。上述结果提示,胃动素可特异性兴奋血管灌流大鼠胃窦收缩运动,该作用通过壁内胆碱能神经介导。     

Induction of specific anti-actin antibodies and their measurement by ELISA technique.

Using the ELISA technique we have been able to quantify antibodies directed against actin and to follow the kinetics of antibody production. Specific anti-actin antisera have been raised in rabbits by immunization with chemically modified white muscle rabbit actin. Two or three dinitrophenyl groups linked per actin molecule were sufficient to break natural tolerance, while linkage of three phosphorylcholine groups to actin was not.     

Distribution of TT virus genomic groups 1-5 in Brazilian blood donors, HBV carriers, and HIV-1-infected patients

Human isolates of the highly prevalent TT virus (TTV) have been classified into five major genomic groups (1-5). The geographical distribution of the groups throughout the world is not well known. Five different PCR assays were developed in an attempt to amplify specifically TTV DNAs of each genomic group. Serum samples collected from 72 Brazilian adults (24 voluntary blood donors, 24 hepatitis B virus (HBV) carriers, and 24 human immunodeficiency virus type 1 (HIV-1)-infected patients) were tested. TTV DNA from at least one genomic group was detected in 11 (46%) blood donors, 13 (54%) HBV carriers, and 24 (100%) HIV-1 patients. All five genomic groups were detected in the three populations, with the exception of group 2 in blood donors. Some samples, negative with all five specific assays, were positive with the commonly used untranslated region (UTR) PCR system. On the other hand, TTV DNA was detected in some samples by using specific assays but not with the UTR PCR. Mixed infections with 2-5 TTV isolates from different groups were detected in 21% blood donors, 29% HBV carriers, and 71% HIV-1 patients. Fifteen PCR products (three obtained with each assay) were sequenced. Most sequences showed high (>86%) homology with those of TTV isolates belonging to their presumed groups. However, three sequences had low homology with all TTV sequences available from the DNA databanks. In conclusion, TTV isolates belonging to all five known genomic groups circulate in Brazil, and the results suggest the existence of new and as yet uncharacterised major genomic groups.     

Adhesion to fibronectin via alpha4 integrin (CD49d) protects B cells from apoptosis induced by serum deprivation but not via IgM or Fas/Apo-1 receptors

Apoptosis is a regulated event crucial to the development and proliferation of normal and malignant B cells. We have studied the role of signals delivered via alpha4 integrin on apoptosis triggered by three different pathways on these cells. For apoptosis induced by serum deprivation, culturing B cells on the recombinant fibronectin fragment H89, a known ligand for alpha4beta1 integrin, resulted in statistically significant (P < 0.005) higher viability values (68%, 65% and 67%) for Ramos, Nalm-6 and EHEB cells, respectively, than culturing cells on poly lysine (42%, 42% and 48%). An antialpha4 MoAb reverted the protecting effect, thus confirming that it was due specifically to alpha4 engagement. Similarly, cells cultured on FN-III4-5, a recently identified fibronectin region which binds activated alpha4 integrin, also showed statistically significant higher viability than poly lysine cultures. Alpha4 engagement however, did not prevent apoptosis induced on Ramos cells via surface IgM. Adhesion of IM-9 cells, a myeloma cell line carrying functional Fas receptors, to the H89 fragment neither increased cell viability upon triggering apoptosis via Fas when compared to poly lysine. These results indicate that alpha4 signalling may overcome B cell apoptosis induced by the lack of growth factors but does not seem to affect the IgM or Fas apoptotic pathways, thus suggesting different intracellular mechanisms for these processes.