The usefulness of a piezo-micromanipulator in ICSI in humans.
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In all, 58 couples suffering from infertility because of congenital bilateral absence of the vas deferens underwent a total of 67 combined microsurgical epididymal aspiration or testicular sperm extraction (TESE) and in-vitro fertilization (IVF) treatments. The oocytes recovered were inseminated by either the microdroplet IVF technique (N = 20), subzonal insemination (SUZI; n = 10) or intracytoplasmic sperm injection (ICSI; n = 37). Of the ICSI cycles, 12 were performed using spermatozoa obtained by TESE. Fertilization rates for epididymal spermatozoa were significantly higher for SUZI (17.9%, 17/95) and ICSI (34.4%), 137/398) than for microdroplet IVF (5.2%, 18/343) cycles. The proportion of cycles in which fertilization was achieved was higher in the SUZI (80%) and ICSI (95%) cycles than in the IVF cycles (45%). Delivery or an ongoing pregnancy was achieved in one (5%) IVF cycle, two (20%) SUZI cycles and seven (18.95) ICSI cycles. SUZI or ICSI using epididymal or testicular spermatozoa significantly improved the oocyte fertility rate. The ICSI procedure was especially advantageous in patients for whom spermatozoa were obtained from a testicular biopsy.
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A magnetic tweezers setup is used to control both the stretching force and the relative linking number DeltaLk of a palindromic DNA molecule. We show here, in absence of divalent ions, that twisting negatively the molecule while stretching it at approximately 1 pN induces the formation of a cruciform DNA structure. Furthermore, once the cruciform DNA structure is formed, the extrusion of several kilo-base pairs of palindromic DNA sequence is directly and reversibly controlled by varying DeltaLk. Indeed the branch point behaves as a nanomechanical gear that links rotation with translation, a feature related to the helicity of DNA. We obtain experimentally a very good linear relationship between the extension of the molecule and DeltaLk. We use then this experiment to obtain a precise measurement of the pitch of B-DNA in solution: 3.61 +/- 0.03 nm/turn.
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The immunoglobulin VH gene rearrangement in a primary cutaneous, large-cell (centroblastic and immunoblastic) B-cell lymphoma was analyzed using a micromanipulation/single-cell polymerase chain reaction technique. In all single B cells obtained from CD20-stained skin sections that gave a polymerase chain reaction product (eight of 27 in biopsy I), the same VHDJH rearrangement, consisting of DP-54-DIR1-JH3a genes, was detected, with no intraclonal nucleotide diversity. Comparison with the most closely related germline counterpart showed significantly altered complementarity determining gene regions as a result of somatic mutations, suggesting an antigen-driven selection and expansion ofthis particular B-cell clone. Interestingly, in a biopsy obtained from the patient 9 mo later, during disease progression (deep muscle infiltration), the lymphoma cells again contained the same VHDJH gene rearrangement (six of 18 in biopsy II) without any further somatic mutations. Therefore, it is suggested that the cutaneous lymphoma characterized throughout this study descended from postgerminal center B-cells.
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A simple device for holding and moving mechanical tools in the region of a sample being viewed in the scanning electron microscope is described. The unit has a 20:1 mechanical reduction and when fitted with a tungsten carbide dental chisel, it is sufficiently rigid to cut biological hard tissues. Alternately, when fitted with an electro-etched tungsten needle, it can be used, in conjunction with specimen stage controls, to remove individual cells from the surface of soft tissues. Examples of these applications are illustrated.
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