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11.
12.
High throughput parallel analysis of hundreds of patient samples for more than 100 mutations in multiple disease genes 总被引:5,自引:0,他引:5
Shuber AP; Michalowsky LA; Nass GS; Skoletsky J; Hire LM; Kotsopoulos SK; Phipps MF; Barberio DM; Klinger KW 《Human molecular genetics》1997,6(3):337-347
As more mutations are identified in genes of known sequence, there is a
crucial need in the areas of medical genetics and genome analysis for
rapid, accurate and cost-effective methods of mutation detection. We have
developed a multiplex allele-specific diagnostic assay (MASDA) for analysis
of large numbers of samples (> 500) simultaneously for a large number of
known mutations (> 100) in a single assay. MASDA utilizes
oligonucleotide hybridization to interrogate DNA sequences. Multiplex DNA
samples are immobilized on a solid support and a single hybridization is
performed with a pool of allele-specific oligonucleotide (ASO) probes. Any
probes complementary to specific mutations present in a given sample are in
effect affinity purified from the pool by the target DNA. Sequence-specific
band patterns (fingerprints), generated by chemical or enzymatic sequencing
of the bound ASO(s), easily identify the specific mutation(s). Using this
design, in a single diagnostic assay, we tested samples for 66 cystic
fibrosis (CF) mutations, 14 beta-thalassemia mutations, two sickle cell
anemia (SCA) mutations, three Tay-Sachs mutations, eight Gaucher mutations,
four mutations in Canavan disease, four mutations in Fanconi anemia, and
five mutations in BRCA1. Each mutation was correctly identified. Finally,
in a blinded study of 106 of these mutations in > 500 patients, all
mutations were properly identified. There were no false positives or false
negatives. The MASDA assay is capable of detecting point mutations as well
as small insertion or deletion mutations. This technology is amenable to
automation and is suitable for immediate utilization for high-throughput
genetic diagnostics in clinical and research laboratories.
相似文献
13.
Dou Q; Tarnuzzer RW; Williams RS; Schultz GS; Chegini N 《Molecular human reproduction》1997,3(11):1005-1014
14.
Rodgers KE; Girgis W; St Amand K; Campeau J; diZerega GS 《Human reproduction (Oxford, England)》1998,13(9):2443-2451
Adhesion formation is a major source of postoperative morbidity and
mortality. In this study, the ability of a variety of lazaroid formulations
[the antioxidant 21-aminosteroid PNU74006F (tirilazad) and the
non-steroidal 2-methylaminochroman derivative PNU83,836E] to reduce i.p.
adhesion formation in three rabbit models was examined. In initial studies,
PNU83836E was administered via Alzet miniosmotic pump to the site of
injury. In the sidewall and double uterine horn models, PNU83,836E was
administered via Alzet miniosmotic pump for the entire postoperative
interval. In the sidewall model, there was a dose- dependent reduction in
the area of the sidewall injury that was involved in adhesions. In the
double uterine horn model, PNU83,836E was administered via Alzet
miniosmotic pump to the area of injury for 1, 2, 3 or 7 days.
Administration for as little as 24 h after surgery significantly reduced
the extent of adhesion formation and the reduction was increased if it was
administered for longer. Further studies were conducted in which various
lazaroid formulations were administered as a bolus at the end of surgery.
In both the sidewall and double uterine horn models, administration of
either PNU83,386E (in citrate buffer) or PNU74006F (in cyclodextrin or
lipid emulsion vehicles) at the end of surgery reduced adhesion formation.
Administration of a bolus of PNU74006F 10 min prior to initiation of
surgery with or without additional treatment at the end of surgery further
increased its efficacy in the reduction of adhesion formation.
Administration of a minimum of 1.5 mg before and after surgery (3 mg total)
was required for maximal efficacy. These studies demonstrate that pre- and
postoperative administration of either a steroidal (PNU74006F) or
non-steroidal (PNU83,836E) lazaroid intraperitoneally reduced the formation
and reformation of postoperative adhesions in three animal models.
相似文献
15.
Combined assessment of intestinal disaccharidases in congenital asucrasia by differential urinary disaccharide excretion. 下载免费PDF全文
Investigation of intestinal disaccharide hydrolysis and permeability by means of a non-invasive differential sugar absorption test was performed in a family containing two siblings with primary sucrase-isomaltase deficiency. The procedure, which depends on measurement of urinary excretion ratios after the oral administration of lactose, sucrose, palatinose, lactulose and L-rhamnose, is capable of simultaneous determination of intestinal lactase, sucrase, and isomaltase activity and lactulose:rhamnose permeability. The results corresponded well with those of disaccharidase assay and histological findings in jejunal biopsy tissue obtained from the patients. Palatinose proved a satisfactory substrate for in vivo assessment of intestinal isomaltase activity. The method described provides a reliable and comprehensive assessment of intestinal disaccharide hydrolysis, and simultaneous estimation of permeability assists discrimination of primary from secondary disaccharidase deficiency. The ability to assess three different disaccharidase activities in addition to intestinal permeability by means of a single test, and the simplicity of preservation and transport of urine samples for sugar analysis, makes this a convenient, definitive method for the investigation of defective sugar absorption in both patients and population groups. 相似文献
16.
Casley William L.; Menzies Allan; Girard Michel; Larocque Lyse; Mousseau Nicole; Whitehouse Larry W.; Moon Thomas W. 《Toxicological sciences》1997,40(2):228-237
The 3-demethylation of caffeine can be used as an index of cytochromeP450 CYP1A2 activity in vivo. We compared the plasma levelsof caffeine and the 3-demethylated metabolite, 1,7-dimethylxanthine,in six common inbred strains (A/J, P/J, BALB/cJ, C3H/HeJ, AKR/J,and SWR/J) and one inbred strain (APN) derived in our laboratoryfrom outbred Swiss-Webster mice on the basis of its relativesusceptibility to acetaminophen-induced hepatotoxicity. We foundsignificant variations between a number of the common strains,all of which produced significantly higher caffeine 3-demethylationindices than our APN strain. In three of the six common strains,there was a significant difference between males and females,with the females having consistently lower 1,7-xanthine/caffeineratios. Hepatic Cyp1a2 expression was compared between APN andC3H/HeJ males. Microsomal methoxyresorufin O-demethylation,acetanilide 4-hydroxylation, and CYP1A2 immunoreactive proteinlevels were significantly higher in C3H/HeJ relative to APNmice, as were hepatic CYP1A2 mRNA levels. These results indicatethe importance of strain and gender to the outcome of pharmacologicalor toxicological studies involving CYP1A2-mediated metabolism,as well as the suitability of the plasma 1,7-dimethylxanthine/caffeineratio as a marker of CYP1A2 activity in the mouse. The strikingdifferences observed between the APN and C3H/HeJ mice suggestthat these strains may be suitable for a genetic analysis ofthe regulation of the basal expression of CYP1A2, a key enzymeIn procarcinogen activation. 相似文献
17.
Renal transplantation has become a treatment of choice for patients with end stage renal disease. A successful transplant is the result of a combination of several factors acting synergistically, such as the degree of HLA compatibility between donor and the recipient, pretransplant blood transfusions, the recipient''s state of immunoreactivity and sensitization, immunosuppressive therapy given in post operative period etc. Donor selection appears to be the most critical factor for the long term success of the organ graft. In this brief review, some of the important parameters of donor selection in renal transplantation are highlighted.KEY WORDS: Histocompatibility (HLA) matching, Cross match, Sensitization 相似文献
18.
Changes in iron histochemistry after hypoxic-ischemic brain injury in the neonatal rat 总被引:6,自引:0,他引:6
Palmer C Menzies SL Roberts RL Pavlick G Connor JR 《Journal of neuroscience research》1999,56(1):60-71
Iron can contribute to hypoxic-ischemic brain damage by catalyzing the formation of free radicals. The immature brain has high iron levels and limited antioxidant defenses. The objective of this study was to describe the early alterations in nonheme iron histochemistry following a hypoxic-ischemic (HI) insult to the brain of neonatal rats. We induced a HI insult to the right cerebral hemisphere in groups of 7-day-old rats. Rats were anesthetized, then their brains were perfused and fixed at 0, 1, 4, 8, 24 hr, and 1, 2, and 3 weeks of recovery. Forty-micron-thick frozen sections were stained for iron using the intensified Perls stain. Increased iron staining was first detected within the cytoplasm of cells with pyknotic nuclei at 4 hr of recovery. Staining increased rapidly over the first 24 hr in regions of ischemic injury. By 7 days recovery, reactive glia and cortical blood vessels also stained. Increased staining in gray matter persisted at 3 weeks of recovery, whereas white matter tracts had fewer iron-positive cells compared to normal. The early increase in iron staining could be caused by an accumulation of iron posthypoxicischemic injury or a change in iron from nonstainable heme iron to stainable nonheme iron. Regardless of the source, our results indicate that there is an increase in iron available to promote oxidant stress in the neonatal rat brain following hypoxia-ischemia. 相似文献
19.
Emergency and elective embolotherapy of various systemic arteries in 64 patients was carried out at a tertiary centre of Armed Forces. Specific indications were haemoptysis (n=43), preoperative (n=18), haematuria (n=1), epistaxis (n=1) and chemoembolization (n=1). The procedures were performed with gelfoam pellets (n=46), gelfoam pellets and absolute alcohol (n=1), polyvinyl alcohol particles (PVA) (n=14), steel coils (n=2) and Adriamycin-in-oil emulsion (n=1). Embolotherapy resulted in complete haemostasis in 37 (82.2%) out of 45 cases of haemorrhage. In eight cases (17.8%), it resulted in significant improvement. Complete haemostasis was achieved in both cases of haematuria and epistaxis. Pre-operative embolotherapy resulted in considerable reduction of peroperative blood loss in all the cases. Chemoembolization of Hepatocellular carcinoma resulted in partial regression of the tumour. The purpose of this study was to assess the efficacy, safety and reliability of vascular embolotherapy for control of life threatening haemorrhage and preoperative reduction of lesions.KEY WORDS: Embolization, Embolotherapy, Haemorrhage 相似文献
20.