Classical t(11;14)(q13;q32) involving IGH-CCND1 is typically associated with aggressive CD5-positive mantle cell lymphoma (MCL). Recently, we identified the IGK variant of this translocation, t(2;11)(p11;q13), in three patients with a leukemic smallcell B-non-Hodgkin lymphoma. In all cases, rearrangements of the IGK and CCND1 genes were demonstrated by fluorescence in situ hybridization. Moreover, we mapped the 11q13 breakpoint of this variant translocation in the 3 0 region of CCND1 which contrasts with the 5 0 breakpoints in a standard t(11;14)(q13;q32). Expression of cyclin D1 was shown in two cases analyzed either at diagnosis or during disease progression. All three patients were asymptomatic at presentation and no initial therapy was required. One patient died of a progressive disease 58 months from diagnosis, and two patients showed stable disease after 12 months of follow-up. In two analyzed cases, mutated IGVH genes were identified. Our findings indicate that variant t(2;11)(p11;q13) does not typify a classical MCL but possibly a more indolent leukemic lymphoma originating from an antigen experienced (mutated) B cell.
We present a sero-epidemiological study of 190 Belgian cases of Lyme borreliosis, a multisystemic disease caused by the spirochaete Borrelia burgdorferi and transmitted by a tick bite. The whole spectrum of clinical pictures was observed in these patients, including "erythema chronicum migrans" (63%), neurological involvement (47%) and arthritis (22%), frequently in combination. Our results are compared to findings in other countries. Among the 437 Ixodes ricinus ticks collected in the Sambre and Meuse valleys around Namur, we discovered 43 ticks (9.8%) with spirochaetes in the midgut. The ecology of these arthropods explains why this infection is more prevalent in the spring and the summer. Perhaps for the same reason, the incidence ranges from low near the coast to medium in the central part and high in the wooded south-eastern part of Belgium. The main conclusions are that Lyme borreliosis is endemic in Belgium, that all the clinical pictures can be observed ant that a clear epidemiological case-definition is needed, combining clinical signs and serological results.
1In melanoma, the lymphocytic infiltrate is a prognostic parameter classified morphologically into "brisk", 2 "non-brisk" and "absent" entailing a functional association that has never been proved. Recently, it has been 3shown that lymphocytic populations can be very heterogeneous, and that anti-PD-1 immunotherapy 4 supports activated T cells. Here, we characterize the immune landscape in primary melanoma by high-5dimensional single cell multiplex analysis in tissue sections (MILAN technique) followed by image analysis, 6RT-PCR and shotgun proteomics. We observed that the brisk and non-brisk patterns are heterogeneous 7 functional categories that can be further sub-classified into active, transitional or exhausted. The 8 classification of primary melanomas based on the functional paradigm also shows correlation with 9 spontaneous regression, and an improved prognostic value than that of the brisk classification. Finally, the 10 main inflammatory cell subpopulations that are present in the microenvironment associated with activation 11 and exhaustion and their spatial relationships are described using neighbourhood analysis. 12
(What’s new in protocol Version 5: an expanded troubleshooting section, more validated antibodies)Multiplexing, labeling for multiple immunostains the very same cell or tissue section in situ, is of considerable interest. The major obstacles to the diffusion of this technique are high costs in custom antibodies and instruments, low throughput, scarcity of specialized skills or facilities. We have validated and detail here a method based on common primary and secondary antibodies, diffusely available fluorescent image scanners and routinely processed tissue sections \(FFPE). It entails rounds of four-color indirect immunofluorescence, image acquisition and removal \(stripping) of the antibodies, before another stain is applied. The images are digitally registered and the autofluorescence is subtracted. Removal of antibodies is accomplished by disulphide cleavage. In excess of 50 different antibody stains can be applied to one single section from routinely fixed and embedded tissue. This method requires a modest investment in hardware and materials and uses freeware image analysis software.
Multiplexing, labeling for multiple immunostains the very same cell or tissue section in situ, is of considerable interest. The major obstacles to the diffusion of this technique are high costs in custom antibodies and instruments, low throughput, scarcity of specialized skills or facilities. We have validated and detail here a method based on common primary and secondary antibodies, diffusely available fluorescent image scanners and routinely processed tissue sections \(FFPE). It entails rounds of four-color indirect immunofluorescence, image acquisition and removal \(stripping) of the antibodies, before another stain is applied. The images are digitally registered and the autofluorescence is subtracted. Removal of antibodies is accomplished by disulphide cleavage. In excess of 50 different antibody stains can be applied to one single section from routinely fixed and embedded tissue. This method requires a modest investment in hardware and materials and uses freeware image analysis software.
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