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Clonal rearrangements in childhood and adult precursor B acute lymphoblastic leukemia: a comparative polymerase chain reaction study using multiple sets of primers.

Ig heavy chain (IgH) and T-cell receptor (TCR) gene rearrangements were investigated by polymerase chain reaction (PCR) amplification of diagnostic tumour samples from 91 patients (57 children and 34 adults, with cut-off at age 16) with precursor B acute lymphoblastic leukemia (ALL). Using primers directed to the framework regions (FR) 1, 2 and 3 of the IgH gene, clonal IgH rearrangements were observed in 82, 58 and 58%, respectively, whereas clonality was presented in 45 and 27% using primers hybridising to the TCR delta and gamma genes. A combination of all five primer sets used resulted in …

Polymerase chain reaction characterization of trypanosomes in Glossina morsitans submorsitans and G. tachinoides collected on the game ranch of Nazinga, Burkina Faso.

The polymerase chain reaction was used to characterize the trypanosomes infecting Glossina morsitans submorsitans and G. tachinoides in the game ranch of Nazinga, Burkina Faso, situated near an agropastoral zone. Dissection of 435 tsetse flies, and PCR analysis of 166 infected flies were conducted to assess the epidemiological situation. Trypanosomes of the Nannomonas subgenus were the most abundant in the two tsetse species (80.4% and 73.7% of identified infections in G. m. submorsitans and G. tachinoides respectively). T. vivax and T. brucei infection rates were comparable between the two ts…

[Detection of viral hemorrhagic septicemia virus (VHSV) in rainbow trout (Oncorhynchus mykiss) by reverse transcription followed by polymerase chain reaction. Diagnostic validation].

Viral haemorrhagic septicaemia virus (VHSV) is a fish pathogen that attacks rainbow trout (Oncorhynchus mykiss) farming. The only diagnostic method recognised by the European community for VHS is based on the detection of the viral agent in cell culture followed by an immunological identification of the pathogen. Reverse transcription followed by a double amplification of the polymerase chain reaction (RT-PCR) for the gene encoding the viral glycoprotein G is proposed here as an alternative method to the virus assay in cell culture. The RT-PCR was found to have three advantages over the viral …

Identification of progesterone-dependent messenger ribonucleic acid regulatory patterns in the rhesus monkey endometrium by differential-display reverse transcription-polymerase chain reaction.

We used differential-display reverse transcription-polymerase chain reaction (DDRT-PCR) to identify different patterns of progesterone (P4)-dependent gene regulation in rhesus monkey endometria. Complementary DNA populations representing the proliferative phase (estrogen dominant, EcDNA) and an inadequate secretory phase (low level of P4, IcDNA) were compared with a cDNA population representing an adequate secretory phase (normal level of P4, PcDNA). We were able to distinguish four different levels of mRNA regulation: 1) up-regulation by P4 during an adequate secretory phase, 2) autologous do…

Detection and localisation of HIV-1 DNA and RNA in fixed adult AIDS brain by polymerase chain reaction/in situ hybridisation technique.

In the brain of patients with AIDS, HIV-1 is localised in a productive form in mononuclear cells. One issue that still needs clarification is whether HIV is localised in cells other than those of mononuclear lineage. Gene amplification by polymerase chain reaction/in situ hybridisation (PCR-IS) could shed light on it. In this study, formalin-fixed, paraffin-embedded brain tissue from ten adult AIDS sufferers was used. Five of them showed evidence of HIV encephalitis (HIVE), five did not show any abnormality. Nested PCR revealed HIV-1 DNA in all HIVE cases and in three of the group without HIVE…

Quantitation of 8 human herpesviruses in peripheral blood of human immunodeficiency virus-infected patients and healthy blood donors by polymerase chain reaction.

Human herpesviruses are associated with morbidity and mortality in persons with compromised immune systems, including patients infected with human immunodeficiency virus (HIV). To investigate the basis for this association, the levels of all 8 human herpesviruses (herpes simplex virus, types 1 and 2, varicella-zoster virus, cytomegalovirus, Epstein-Barr virus, and human herpesvirus 6, human herpesvirus 7, and human herpesvirus 8) were measured with the quantitative polymerase chain reaction (PCR). Viral DNA was measured in the whole blood of 20 HIV-infected patients and compared with levels in…

Clonal integration and expression of human T-cell lymphotropic virus type I in carriers detected by polymerase chain reaction and inverse PCR.

Adult T-cell leukemia (ATL) is a neoplasm of mature helper (CD4) T lymphocytes, and human T-cell lymphotropic virus type-I (HTLV-I) has been suggested to be the causative virus of ATL. HTLV-I integrates its proviruses into random sites in host chromosomal DNA. Clonal integration has been observed in patients with ATL, including smoldering, chronic, and acute states. However, random and/or polyclonal integration has only been reported in a few asymptomatic HTLV-I carriers. To clarify the clonality of HTLV-I-infected cells in carriers, we used an inverse polymerase chain reaction (IPCR), which i…

Характеристика геномной РНК коксакевируса B3 в миокардите миокарда: надежность прямой последовательности продуктов цепной реакции с обратной транскрипцией полимеразы.

Мыши SWR развивают вирусный миокардит гистологическим образом, похожий на болезнь человека, после прививки кардиовирулентом Коксакивирусом B3 (CVB3), восстановленным из секвенированного клона CDNA штамма Нэнси. Последовательность из 215 нуклеотидов, или 628 нуклеотидов в типичных случаях, 5'непереводимой области (5'NTR) генома CVB3 была увеличена из миокардных образцов инфицированных мышей путем обратной транскрипции полимерной реакции (RT-NPCR). Для проверки последовательности вирусных нуклеотидов и обнаружения частотности мутации вирусных РНК последовательность нуклеотидов продуктов ПНКПР оп

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SWR mice develop viral myocarditis histologically similar to the human disease following inoculation with a cardiovirulent Coxsackievirus B3 … 3 genome was amplified from myocardial samples of the infected mice by reverse transcription-nested polymerase chain reaction (RT-NPCR). In order to verify the viral nucleotide sequence and detect the mutation frequency of the viral RNA, the nucleotide sequence of NPCR products were determined by direct sequencing in both orientations. The amplified products from mouse hea …

Rapid polymerase chain reaction-based detection of the causative agent of cat scratch disease (Bartonella henselae) in formalin-fixed, paraffin-embedded samples.

Bartonella (formerly Rochalimaea) henselae (Bh) plays a central role in cat scratch disease. A polymerase chain reaction (PCR)-based assay that can detect Bh DNA in formalin-fixed, paraffin-embedded (FF-PE) samples would have utility in the evaluation of processed lymph nodes suggestive of this disorder. Fresh or FF-PE cultures of Bh and related species were analyzed. Thirteen lymph nodes (12 FF-PE and one fresh cell suspension) with necrotizing suppurative granulomatous inflammation and seven FF-PE negative control lymph nodes were also evaluated. PCR was performed using a novel, hemi-nested …

Sleeping sickness in Zaire: a nested polymerase chain reaction improves the identification of Trypanosoma (Trypanozoon) brucei gambiense by specific kinetoplast DNA probes.

Blood samples collected in the sleeping sickness focus of Boma, Zaire, from human patients and domestic animals were analysed by polymerase chain reaction (PCR) for the presence of trypanosome DNA. The comparison of PCR and miniature anion exchange centrifugation technique (m-AECT) results clearly shows that in domestic animals mixed infections (Trypanozoon/Trypanosoma [Nannomonas] congolense) are more frequently diagnosed by PCR than by m-AECT. Trypanozoon positive blood samples were further analysed for Trypanosoma (Trypanozoon) brucei gambiense. For that purpose amplified minicircle kinetop…

Genotyping human and bovine isolates of Cryptosporidium parvum by polymerase chain reaction-restriction fragment length polymorphism analysis of a repetitive DNA sequence.

In order to define transmission routes of cryptosporidiosis and develop markers that distinguish Cryptosporidium parvum isolates, we have identified 2 polymorphic restriction enzyme sites in a C. parvum repetitive DNA sequence. The target sequence was amplified by polymerase chain reaction from 100 to 500 oocysts and the amplified product was subjected to restriction enzyme digestion. Typing of 23 isolates showed that 10/10 calf isolates had the same profile. In contrast, 2 patterns were observed among human isolates: 7/13 displayed the calf profile, and 6/13 presented another pattern. The PCR…

Detection of common viruses using the polymerase chain reaction to assess levels of viral presence in type 1 (insulin-dependent) diabetic patients.

The polymerase chain reaction was used to detect a range of common viruses in the peripheral blood of Type 1 diabetic and non-diabetic control patients in order to identify any abnormal viral presence, with possible roles in the pathogenesis of Type 1 diabetes. Peripheral blood from 17 newly diagnosed Type 1 diabetic patients, 38 Type 1 diabetic patients with disease of longer duration, and 43 age and sex matched non-diabetic controls was obtained. Samples were screened for cytomegalovirus, Epstein-Barr virus, enterovirus (including coxsackie), and mumps virus. Cytomegalovirus was detected in …

Sensitive and specific colorimetric dot assay to detect eastern equine encephalomyelitis viral RNA in mosquitoes (Diptera: Culicidae) after polymerase chain reaction amplification.

A sensitive and specific colorimetric dot assay following polymerase chain reaction (PCR) method has been developed to detect 0.1 pg of eastern equine encephalomyelitis viral (EEEV) RNA. The assay is 250-fold more sensitive than analysis by electrophoresis and is based on converting a 291-nucleotide sequence of the viral coat protein amino terminus into a double-stranded DNA (dsDNA) and amplifying the DNA using a specific primer pair and PCR. The amplified complementary DNA (cDNA) is denatured adsorbed onto a nylon strip, baked, and detected with a digoxigenin-labeled probe. Dots with viral cD…

Detection of placental growth hormone variant and chorionic somatomammotropin ribonucleic acid expression in human trophoblastic neoplasms by reverse transcriptase-polymerase chain reaction.

Attempts to assess human placental GH variant (hGH-V) and chorionic somatomammotropin (hCS) RNA in choriocarcinoma cell lines have been hampered by low levels of expression and limited sensitivity of RNA blotting analysis. We examined human choriocarcinoma BeWo, JAR, and JEG-3 cell lines as well as samples of complete hydatidiform moles for expression of members of the human GH (hGH) gene family using reverse transcriptase-polymerase chain reaction. A single and common set of primers was designed and used to detect products of the hGH/hCS genes as well as distinguish processed RNA from any con…

Neoplastic hormone-producing cells of the placenta produce and secrete parathyroid hormone-related protein. Studies by immunohistology, immunoassay, and polymerase chain reaction.

BACKGROUND: Parathyroid hormone-related protein (PTHrP) was originally discovered as a product of tumors associated with hypercalcemia. Subsequent studies have demonstrated the widespread production of PTHrP in both fetal and adult tissues, among them the placenta, where it has been identified in amniotic epithelial cells and trophoblasts. However, studies of PTHrP production by the neoplastic placenta have not been reported. EXPERIMENTAL DESIGN: We have studied placental cells and tissues for PTHrP by immunohistology, immunoassay, and the polymerase chain reaction. We studied normal placenta…

Chlamydia trachomatis-specific antibodies in patients with pelvic inflammatory disease: comparison with isolation in tissue culture or detection with polymerase chain reaction.

OBJECTIVE: The detection of acute phase antibodies against C trachomatis and its comparison with tissue culture or polymerase chain reaction (PCR) on samples of cervix and urethra obtained from patients with pelvic inflammatory disease (PID). METHODS: In the academic hospital Dijkzigt, Rotterdam, The Netherlands, prospective investigations were performed on 49 consecutive patients who were admitted with the diagnosis of PID. Infections with C trachomatis were traced using tissue culture, PCR or by determining acute phase IgG and IgM antibodies. Differences between the sensitivities of serolog…

Use of arbitrarily primed polymerase chain reaction analysis to type disease and carrier strains of Neisseria meningitidis isolated during a university outbreak.

Disease and carrier isolates of Neisseria meningitidis from two regions of the United States were typed by the arbitrarily primed polymerase chain reaction (PCR), or random amplified polymorphic DNA (RAPD), method. This technique generates strain-specific arrays of amplified DNA fragments using low-stringency PCR with single, arbitrarily chosen primers. Each of 3 disease isolates and 7 of 11 carrier isolates from an outbreak at the University of Connecticut were indistinguishable using each of 4 primers. In contrast, 22 other isolates (the remaining 4 carrier isolates plus 18 disease and carri…

Detection of monoclonality in low-grade B-cell lymphomas using the polymerase chain reaction is dependent on primer selection and lymphoma type.

Detection of B-cell monoclonality using the polymerase chain reaction (PCR) promises the quick and cost-effective separation of monoclonal from polyclonal B-cell disease. However, the efficiency of the method has yet to be fully assessed, particularly with regard to disease type and selection of PCR primers. We have evaluated two approaches based on amplification of the immunoglobulin heavy chain gene using framework 2 (Fr2) and framework 3 (Fr3) region primers. Frozen tissue samples from 94 cases of low-grade B-cell lymphoma were investigated, all of which had previously been shown to be mono…

Clinical significance of bcr-abl gene rearrangement detected by the polymerase chain reaction after allogeneic bone marrow transplantation in chronic myelogenous leukemia.

Although serial detection of bcr-abl positive cells by PCR appears able to identify distinct patient groups with different risks of relapse following BMT, there remain many unanswered questions regarding the clinical utility and biological significance of PCR detectable cells in this disease. Many of the studies summarized have conflicting results and the influence of various clinical parameters which are known to affect the risk of relapse post-BMT has not yet been consistently associated with the ability to detect bcr-abl positive cells by PCR. These clinical parameters include GVHD, T-cell

Detection of human T-cell lymphotropic virus type 1 infection by the polymerase chain reaction using dried blood specimens on filter papers.

A simple method for detection of proviral DNA sequences of human T-cell lymphotropic virus type 1 (HTLV-1) was developed using dried blood specimens on filter papers. The whole blood was blotted onto the Guthrie paper. After the blood has dried, the blotted paper was punched out into small discs. The discs were then boiled to prepare the template for PCR (filter paper-PCR method). The filter paper-PCR method detected even a single HTLV-1-infected cell in three discs. The sensitivity of the filter paper-PCR method was equivalent to that of the method in which DNA was extracted with phenol and u