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Melanoma is an immunogenic tumour. The development of skin depigmentation or melanoma-associated leucoderma (MAL) has been associated with favourable clinical outcome in patients with metastatic melanoma, especially after immunotherapy. Evidence for clinically meaningful enhancement of melanoma-directed autoimmunity, as indicated by MAL, after radiotherapy without immunotherapy has not yet been published.We investigated whether a patient with stage IV melanoma, who developed leucoderma in the irradiated skin areas following radiotherapy and experienced exceptional disease-free survival of 3 years despite brain metastasis, possessed antimelanoma immunity that could be linked to the favourable disease course.A detailed immunological analysis was performed consisting of immunohistochemistry of several melanoma tissues, and analyses of T cells isolated from the blood and MAL skin tissue for melanocyte/melanoma specificity and functionality, as well as the presence of a melanoma-specific antibody response.Immunological analyses showed the presence of CD8+ T cells and antibody responses directed against melanocyte differentiation antigens expressed in the primary tumour, lymph node and brain metastasis, indicating adequate tumour recognition by activated T cells.The immune responses found in this patient, probably enhanced by radiotherapy, are thought to have contributed to his favourable clinical course. Radiotherapy may act as local immunotherapy in patients with melanoma by destroying melanocytes, leading to the induction, or enhancement, of already existent antimelanoma immunity. As in patients treated with immunotherapy, this may lead to MAL, also at distant sites from the treated area. This patient is a clear example of the positive prognostic value of MAL, which is possibly induced by radiotherapy, for patients with melanoma.

In a man aged 56 years with dysphagia, an oesophageal biopsy was found to contain an adenocarcinoma. In view of the discrepancy between the clinical and the histological pictures, the biopsy was repeated; in a second biopsy no carcinoma was demonstrable. DNA microsatellite analysis proved that the first biopsy originated from another person. The scheduled oesophageal resection was cancelled and the patient was reassured. A woman aged 77 years underwent gastrectomy because of biopsy samples with adenocarcinoma. However, no tumour was found in the resected stomach. DNA microsatellite analysis showed that the biopsy samples indeed originated from the patient. Unfortunately, mix-up of patient specimens occasionally occurs. Especially a discrepancy between the clinical picture and the histological diagnosis must raise the suspicion of a specimen mix-up. In such cases, DNA microsatellite analysis can give a rapid and reliable answer whether a mix-up has indeed taken place.

The B cell Ag-receptor complex is composed of membrane immunoglobulin (mIg) and the mb-1/B29 heterodimer. In order to obtain insight into the architecture of the B cell receptor complex, we have looked for conditions that disrupt all disulfide bridges in the complex without affecting the noncovalent interaction between the mIg heavy chain and one or both members of the associated heterodimer. We show that in the presence of the reducing agent beta-mercaptoethanol the m mu, m delta, and m gamma heavy chains remain selectively associated with the B29 members. Our findings implied that if isotype-related differences exist between the mIg-associated dimers, they may reside in B29 and not, as initially suggested, in mb-1. However, sequence analyses of B29 gene transcripts from B cells expressing mIgM, mIgD, or mIgG only revealed no differences in their nucleotide composition. Thus, in spite of their close physical interaction with mIg heavy chain classes, which are significantly distinct in the C-terminal regions, no isotype-specific forms of B29 seem to exist.

The diagnosis severe combined immunodeficiency was made in a male infant at the age of 18 wk. Known causes of severe combined immunodeficiency were excluded. The activity of total 5'-nucleotidase (E.C. 3.1.3.5) in the PBMC was found to be strongly decreased. Analysis of the peripheral blood revealed a lymphocytosis, mainly of CD8+ T cells. These lymphocytes expressed high levels of CD29, CD38, CD45RA, and MHC class II molecules but no CD25, CD26, CD27, or CD28 Ag. The cells proliferated poorly to all T cell stimulants tested and no helper activity for IgM secretion could be induced. In contrast to the poor proliferative responses, high levels of TCR-induced cytolytic activity, without lymphokine-activated killer-cell outgrowth, were induced by CD3 mAb. Analysis of TCR-beta gene rearrangements indicated that two clonal populations constituted the majority of the E-rosette+ peripheral blood fraction. Moreover, the vast majority of the CD8+ cells were found to react with a mAb to V beta 3. Polymerase chain reaction on cDNA from peripheral blood cells with primers that amplify TCR V beta elements showed, in agreement with the fluorescence data, an overrepresentation of V beta 3 but absence of usage of approximately 50% of the other V beta elements. Thus, in a severe combined immunodeficiency patient, CD8+ T cells with limited T cell receptor usage and restricted effector functions were found. The observed alterations in the 5'-nucleotidase levels may be secondary to the outgrowth of this population.

Recent evidence indicates that the transmembrane form of IgM on murine and human B lymphocytes is physically associated with at least two proteins, forming a disulfide-linked dimer, which may control cell surface expression of IgM and also play a role in signal transduction after Ag binding (by analogy with the TCR-associated CD3 components in T lymphocytes). We have used mAb and polyclonal antibodies against an intracytoplasmic epitope on one of these polypeptides (previously identified in murine B cells as the product of the B cell specific mb-1 gene) to study the distribution of the IgM-associated dimer in human cells. By immunocytochemical staining of normal and neoplastic B cells, we show that the human mb-1 protein appears early in B cell differentiation, probably before expression of cytoplasmic mu-chain, and persists until the plasma cell stage, where it is seen as an intracytoplasmic component. According to immunohistologic analysis of reactive lymphoid tissue and lymphoma samples, mb-1 protein is completely B cell specific. Anti-mb-1 also labels B cell areas in tissues from seven different mammalian species. Finally, the Ig-associated dimer could be isolated from human hairy-cell leukemia cells in high purity and yield by affinity chromatography using anti-mb-1 antibody. Mice immunized with this material have produced a strong polyclonal response, so that it should now be possible to prepare a panel of new mAb reactive with different epitopes on both mb-1 and on its associated polypeptide(s).

7/8embrane IgM (mIgM) on human B lymphocytes is noncovalently associated with a disulfide-linked dimer that contains phosphoproteins of 47 and 37 kDa. In this study, the biochemical properties and the identity of these Ag receptor-associated components have been addressed. Both subunits carry N-linked carbohydrate groups. After deglycosylation, the 47-kDa and 37-kDa proteins have similar molecular masses, of about 23 kDa, and relatively acidic but different isoelectric points. The accumulated data, together with a previously performed comparison of tryptic peptides, suggest that the two components are structurally distinct and possibly encoded by different genes. Indeed, a mAb, raised against a synthetic peptide that was made on the basis of the published carboxyl-terminal amino acid sequence of the human mb-1 gene product, specifically reacted with the 47-kDa but not the 37-kDa subunit. None of the established B cell-specific mAb characterized in the Fourth International Workshop on Leukocyte Antigens, including CD24, CD37, and CD72, detect the mIgM-linked heterodimer, which makes it a newly defined human B cell Ag.

7/8embrane IgM (mIgM) on human B lymphocytes is noncovalently associated with a disulfide-linked dimer that contains phosphoproteins of 47 and 37 kDa. In this study, the biochemical properties and the identity of these Ag receptor-associated components have been addressed. Both subunits carry N-linked carbohydrate groups. After deglycosylation, the 47-kDa and 37-kDa proteins have similar molecular masses, of about 23 kDa, and relatively acidic but different isoelectric points. The accumulated data, together with a previously performed comparison of tryptic peptides, suggest that the two components are structurally distinct and possibly encoded by different genes. Indeed, a mAb, raised against a synthetic peptide that was made on the basis of the published carboxyl-terminal amino acid sequence of the human mb-1 gene product, specifically reacted with the 47-kDa but not the 37-kDa subunit. None of the established B cell-specific mAb characterized in the Fourth International Workshop on Leukocyte Antigens, including CD24, CD37, and CD72, detect the mIgM-linked heterodimer, which makes it a newly defined human B cell Ag.

B chronic lymphocytic leukemia (B-CLL) are clonal populations of mIgM+ or mIgM+/mIgD+ CD5+ B cells that appear to be arrested in the follicular mantle-zone B-cell stage. Functional analyses have shown two groups of B-CLL that can be distinguished based on their capacity to proliferate in response to B-cell antigen receptor complex (BCR) cross- linking. To investigate the molecular basis for this phenomenon, we have analyzed both architecture and functional properties of BCR complexes on these two groups of B-CLL. Both groups were found to express structurally similar BCR. However, protein tyrosine kinase (PTK) activity associated with and specific for BCR constituents was strongly diminished in nonresponsive B-CLL. Moreover, the PTK-dependent assembly of Shc/Grb2 complexes, which may couple the BCR to p21ras, was absent in these B-CLL. Finally, of all PTKs tested, the expression of PTK syk was found to be considerably lower in nonresponsive B-CLL. Thus, absence of mitogenic responses upon BCR cross-linking in particular B-CLL was found to be strictly correlated with diminished induction of BCR-associated PTK activity and lower levels of PTK syk. Because nonresponsive B-CLL closely resembles tolerant autoreactive B cells both functionally and biochemically, distinction between B-CLL with respect to functional properties in vitro may be determined by differences in antigen encounter in vivo.