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2.
H. L. Zaaijer H.T.M. Cuypers H.W. Reesink J. H. ten Veen S. C. E. Schoon-Visser and P. N. Lelie 《Vox sanguinis》1994,66(1):78-80
Recently, an assay for detection of proviral HIV-1 DNA in leukocytes became commercially available. This assay (Amplicor HIV-1 test, Roche Diagnostic Systems) multiplies HIV-1 DNA up to a detectable level, using the polymerase chain reaction. We studied performance of this assay on 74 samples from HIV-1-infected patients and on 41 samples from healthy blood donors. Twice a negative control sample appeared to be erroneously reactive. However, sensitivity and specificity on the patient and donor samples both were 100%. To avoid false-positive results, we advise to repeat initially reactive samples if no other data confirm HIV-infection. 相似文献
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5.
Storage conditions of blood samples and primer selection affect the yield of cDNA polymerase chain reaction products of hepatitis C virus.
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H T Cuypers D Bresters I N Winkel H W Reesink A J Weiner M Houghton C L van der Poel P N Lelie 《Journal of clinical microbiology》1992,30(12):3220-3224
We have noticed that suboptimal specimen processing and storage conditions may cause false-negative results in the detection of hepatitis C virus (HCV) RNA in plasma or serum. To establish the influence of specimen handling in a serological laboratory on the rate of detection of HCV RNA by the cDNA polymerase chain reaction (cDNA-PCR), we tested routine serum samples and fresh-frozen plasma samples from the same bleeding from confirmed anti-HCV-positive blood donors. When primers from the NS3/NS4 region were used, HCV RNA was detected in fresh-frozen plasma from 67% of the donors, whereas positive results were obtained with only 50% of the serum samples that had been subjected to routine serological procedures. Analysis of the same samples with primers from the highly conserved 5'-terminal region (5'-TR) revealed an HCV RNA detection rate of 92% for both the routine and the fresh-frozen samples. However, the yield of the amplification product in routine samples was strongly reduced compared with that in fresh-frozen plasma. Comparison of both primer sets for cDNA-PCR showed that the 5'-TR primer set was 10- to 100-fold more effective in detecting HCV RNA. We also analyzed the effect of storage of whole EDTA-blood and serum at room temperature and at 4 degrees C on the yield of the amplification product. A rapid decline in detectable HCV RNA of 3 to 4 log units was observed within 14 days when whole blood and serum were stored at room temperature. By contrast, no perceptible reduction in the cDNA-PCR signal was found in freshly prepared serum stored at 4 degrees C. 相似文献
6.
Calcium ionophore-induced acrosome reaction correlates with fertilization rates in vitro in patients with teratozoospermic semen 总被引:5,自引:3,他引:5
The aim of this study was to determine the relationship between calcium
ionophore A23187-induced acrosome reaction (AR) and sperm fertilizing
ability. Semen samples remaining after preparation for standard IVF were
studied in 109 patients who had sperm concentrations > or =20 x
10(6)/ml. Ionophore-induced AR was performed on motile spermatozoa selected
by centrifugation on a Percoll gradient. Semen analysis was performed using
standard methods. Patients with higher (>50%, n = 76) fertilization
rates had significantly higher ionophore-induced AR than patients with
lower (<50%, n = 33) fertilization rates (49 +/- 14 versus 38 +/- 21%, P
< 0.05). When the data from all patients were analysed by logistic
regression, only the percentage sperm motility in insemination medium and
ionophore-induced AR were significantly related to fertilization rates.
Similar results were also obtained when the data from a subgroup of
patients with poor (<15% normal) sperm morphology were analysed.
However, when patients with normal sperm morphology > or =15% were
analysed separately, only sperm count and the percentage of spermatozoa
with progressive motility in semen were significantly related to
fertilization rates. In conclusion, ionophore- induced AR was significantly
related to fertilization rates in vitro mainly in patients with
teratozoospermic semen. Tests for ionophore- induced AR may provide
additional information about sperm fertilizing ability but may not indicate
specific defects of the physiological AR.
相似文献
7.
Detection of reactive oxygen species (ROS) and apoptosis in human fragmented embryos 总被引:8,自引:2,他引:8
Yang HW; Hwang KJ; Kwon HC; Kim HS; Choi KW; Oh KS 《Human reproduction (Oxford, England)》1998,13(4):998-1002
In human in-vitro fertilization (IVF)-embryo transfer, the in-vitro culture
environment differs from in-vivo conditions in that the oxygen
concentration is higher, and in such conditions the mouse embryos show a
higher concentration of reactive oxygen species (ROS) in simple culture
media. ROS are believed to cause damage to cell membranes and DNA
fragmentation in somatic cells. This study was conducted to ascertain the
level of H2O2 concentration within embryos and the morphological features
of cell damage induced by H2O2. A total of 62 human oocytes and embryos (31
fragmented, 15 non-fragmented embryos, 16 unfertilized oocytes) was
obtained from the IVF-embryo transfer programme. The relative intensity of
H2O2 concentrations within embryos was measured using
2',7'-dichlorodihydrofluorescein diacetate by Quanti cell 500 fluorescence
imaging and DNA fragmentation was observed with transmission electron
microscopy and an in-situ apoptosis detection kit. The H2O2 concentrations
were significantly higher in fragmented embryos (72.21 +/- 9.62, mean +/-
SEM) compared to non-fragmented embryos (31.30 +/- 3.50, P < 0.05) and
unfertilized oocytes (30.75 +/- 2.67, P < 0.05). Apoptosis was observed
only in fragmented embryos, and was absent in non-fragmented embryos.
Electron microscopic findings confirmed apoptotic bodies and cytoplasmic
condensation in the fragmented blastomeres. We conclude that there is a
direct relationship between increased H2O2 concentration and apoptosis, and
that further studies should be undertaken to confirm these findings.
相似文献
8.
Stein TP; Oram-Smith JC; Leskiw MJ; Wallace HW; Long LC; Leonard JM 《The American journal of physiology》1976,230(5):1321-1325
9.
Interchromosomal duplications of the adrenoleukodystrophy locus: a phenomenon of pericentromeric plasticity 总被引:13,自引:5,他引:13
Eichler EE; Budarf ML; Rocchi M; Deaven LL; Doggett NA; Baldini A; Nelson DL; Mohrenweiser HW 《Human molecular genetics》1997,6(7):991-1002
A 9.7 kb segment encompassing exons 7-10 of the adrenoleukodystrophy (ALD)
locus of the X chromosome has duplicated to specific locations near the
pericentromeric regions of human chromosomes 2p11,10p11, 16p11 and 22q11.
Comparative sequence analysis reveals 92-96% nucleotide identity,
indicating that the autosomal ALD paralogs arose relatively recently during
the course of higher primate evolution (5-10 million years ago). Analysis
of sequences flanking the duplication region identifies the presence of an
unusual GCTTTTTGC repeat which may be a sequence-specific integration site
for the process of pericentromeric- directed transposition. The breakpoint
sequence and phylogenetic analysis predict a two-step transposition model,
in which a duplication from Xq28 to pericentromeric 2p11 occurred once,
followed by a rapid distribution of a larger duplicon cassette among the
pericentromeric regions. In addition to facilitating more effective
mutation detection among ALD patients, these findings provide further
insight into the molecular basis underlying a pericentromeric-directed
mechanism for non- homologous interchromosomal exchange.
相似文献
10.
E. P. Mauser-Bunschoten G. Roosendaal H. M. van den Berg D. Bresters A. A. J. van Drimmelen H. T. M. Cuypers P. N. Lelie H. W. Reesink C. L. Der Van Poel 《Journal of medical virology》1995,45(3):241-246
Serum samples from 316 patients visiting the Dutch National Hemophilia Center were collected from 1979 to 1993 and stored at ?30°C. Patients were placed into three different groups: (1) patients ever treated with large pool non-hepatitis C virus (HCV)-safe concentrate (n=179); (2) patients treated with cryoprecipitate (n = 125); and (3) patients treated exclusively with HCV-save concentrate (n=12). In order to examine the prevalence of HCV infection in the different treatment groups serum samples were tested retrospectively for anti-HCV antibody using second generation enzyme-linked immunosorbent assay (ELISA) and recombinant immunoblot assay (RIBA-2). Significant differences in the prevalence of HCV infection were found between these 3 groups (group 1: 99%, group 2: 66%, group 3: 0%). The safety of currently administered clotting products is demonstrated in 57 patients who remained without HCV markers between 1989 and 1993. To examine the natural course of HCV infection fresh-frozen plasma samples were obtained recently from a subgroup of 277 hemophilia patients for HCV-RNA detection by a well-validated cDNA-PCR assay. In contrast to other reports, no evidence was found for seronegative HCV carriers. None of 52 patients without anti-HCV had detectable HCV-RNA. Of 225 patients with anti-HCV, 182 (81%) were HCV-RNA positive. None of 39 anti-HCV positive patients with a negative HCV-RNA reaction had serum alanine aminotransferase (ALT) levels above 50 U/l, whereas 44% of HCV-RNA positive patients had persistently elevated ALT levels above 50 U/l. These results indicate that 20% of hemophilia patients who have been infected with HCV in the past eliminated the virus or have viral replication below the detection limit of polymerase chain reaction (PCR) without biochemical evidence of liver damage. © 1995 Wiley-Liss, inc. 相似文献