A nested polymerase chain reaction(N-PCR)for the spegific detection of Helicobacterpylori(H.pylori)was developed with two primer pairs(nested primers)derived from ureasegene A of H.pylori.The N-PCR was used to detect ...A nested polymerase chain reaction(N-PCR)for the spegific detection of Helicobacterpylori(H.pylori)was developed with two primer pairs(nested primers)derived from ureasegene A of H.pylori.The N-PCR was used to detect 21 different samples of H.pylori including20 clinical isolates and 1 reference strain NCTC 14126,but it was negative for other bacterialspecies,showing the N-PCR assay to be 100% specific.Tenfold serial dilution experiments re-vealed the detection of as little as 0.1 fg of H.pylori DNA by N-PCR.To evaluate the PCR as-say for clinical samples,gastric biopsies were tested with N-PCR,and the results were comparedwith those of culture,urease test and histologic examination(reference standard,RS).In 30biopsy specimens,H.pylori DNA sequences were detected by PCR in all of 20(100%)positivetissue and none of the 10 negative tissues.PCR is a specific and sensitive method that can detectthe presence of H.pylori without the need for culture and would have significant importance di-agnostically and epidemiologically.展开更多
Polymerase chain reaction (PCR) was used to detect the presence of Barrelia burgdoferi DNA in biological samples from patients with sarcoidosis. The target DNA sequence was of chromosomal origin. The amplified DNA seq...Polymerase chain reaction (PCR) was used to detect the presence of Barrelia burgdoferi DNA in biological samples from patients with sarcoidosis. The target DNA sequence was of chromosomal origin. The amplified DNA sequence was analyzed by agarose gel electrophoresis, PAGE with silver staining, and the identity of amplified DNA was confirmed by restriction enzyme cleavage and DNA-DNA hybridization with a 32P-labelled probe. The assay was sensitive to fewer than two copies of B. burgdorferi genome, even in the presence of a 104-fold excess of human eukaryotic DNA , and was also specific to different B. burgdorferi strains tested. Sera serologically positive to B. burgdcrferi (n=26), bronchoalveolar lavage fluid and supernatant of BALF (n=26) and peripheral blood (n=9) from sarcoidosis patients were tested. The positive rate was low (4/26, 2/26. and 0/9, respectively). It was considered that DNA from B. burgdor ferimay be identified in a minority of patients with sarcoidosis, and it may play a pathogenetic role in such cases. More studies need to be done before advancing the hypothesis of an etiologic role of B. burgdorferi in sarcoidosis.展开更多
Application of polymerase chain reaction (PCR) to the hypervariable segment of the immunoglobulin heavy chain (IgH) gene allows detection of minimal residual disease (MRD) at a level of one leukemic cell in 103 104 no...Application of polymerase chain reaction (PCR) to the hypervariable segment of the immunoglobulin heavy chain (IgH) gene allows detection of minimal residual disease (MRD) at a level of one leukemic cell in 103 104 normal marrow cells. We used seminested PCR to amplify the DNA fragment of the complementarity-determining region-Ⅲ of the IgH gene from leukemic cell specimens of patients with leukemia. There was IgH gene rearrangement in 25 of 34 ( 74% ) acute lymphohlastic leukemia (ALL) patients, 3 of 4 (75%) chronic lymphocytic leukemia (CLL) patients. Five of 33 (15%) acute myeloid leukemia (AML) patients and 0 of 9 (0%) chronic myeloid leukemia (CML) patients. ALL PCR positive cases were confirmed by Southern blot analysis. Our results indicated that (1) the seminested PCR technique was found to have a higher sensitivity and less false-negative results than onestage PCR; (2) IgH gene rearrangement may not be Iimited to lymphoid leukemia of B cell lineage. In some patients,the leukemic transforming event may involve stem cells capable of both B cell and myeloid differentiation, or AML cells may differentiate along different lineages with the predominant appearance of one or the other subclone in the course of the disease. The mechanism needs to be further investigated.展开更多
文摘A nested polymerase chain reaction(N-PCR)for the spegific detection of Helicobacterpylori(H.pylori)was developed with two primer pairs(nested primers)derived from ureasegene A of H.pylori.The N-PCR was used to detect 21 different samples of H.pylori including20 clinical isolates and 1 reference strain NCTC 14126,but it was negative for other bacterialspecies,showing the N-PCR assay to be 100% specific.Tenfold serial dilution experiments re-vealed the detection of as little as 0.1 fg of H.pylori DNA by N-PCR.To evaluate the PCR as-say for clinical samples,gastric biopsies were tested with N-PCR,and the results were comparedwith those of culture,urease test and histologic examination(reference standard,RS).In 30biopsy specimens,H.pylori DNA sequences were detected by PCR in all of 20(100%)positivetissue and none of the 10 negative tissues.PCR is a specific and sensitive method that can detectthe presence of H.pylori without the need for culture and would have significant importance di-agnostically and epidemiologically.
文摘Polymerase chain reaction (PCR) was used to detect the presence of Barrelia burgdoferi DNA in biological samples from patients with sarcoidosis. The target DNA sequence was of chromosomal origin. The amplified DNA sequence was analyzed by agarose gel electrophoresis, PAGE with silver staining, and the identity of amplified DNA was confirmed by restriction enzyme cleavage and DNA-DNA hybridization with a 32P-labelled probe. The assay was sensitive to fewer than two copies of B. burgdorferi genome, even in the presence of a 104-fold excess of human eukaryotic DNA , and was also specific to different B. burgdorferi strains tested. Sera serologically positive to B. burgdcrferi (n=26), bronchoalveolar lavage fluid and supernatant of BALF (n=26) and peripheral blood (n=9) from sarcoidosis patients were tested. The positive rate was low (4/26, 2/26. and 0/9, respectively). It was considered that DNA from B. burgdor ferimay be identified in a minority of patients with sarcoidosis, and it may play a pathogenetic role in such cases. More studies need to be done before advancing the hypothesis of an etiologic role of B. burgdorferi in sarcoidosis.
文摘Application of polymerase chain reaction (PCR) to the hypervariable segment of the immunoglobulin heavy chain (IgH) gene allows detection of minimal residual disease (MRD) at a level of one leukemic cell in 103 104 normal marrow cells. We used seminested PCR to amplify the DNA fragment of the complementarity-determining region-Ⅲ of the IgH gene from leukemic cell specimens of patients with leukemia. There was IgH gene rearrangement in 25 of 34 ( 74% ) acute lymphohlastic leukemia (ALL) patients, 3 of 4 (75%) chronic lymphocytic leukemia (CLL) patients. Five of 33 (15%) acute myeloid leukemia (AML) patients and 0 of 9 (0%) chronic myeloid leukemia (CML) patients. ALL PCR positive cases were confirmed by Southern blot analysis. Our results indicated that (1) the seminested PCR technique was found to have a higher sensitivity and less false-negative results than onestage PCR; (2) IgH gene rearrangement may not be Iimited to lymphoid leukemia of B cell lineage. In some patients,the leukemic transforming event may involve stem cells capable of both B cell and myeloid differentiation, or AML cells may differentiate along different lineages with the predominant appearance of one or the other subclone in the course of the disease. The mechanism needs to be further investigated.
