Cytomegalovirus Monitoring by Polymerase Chain Reaction of Whole Blood Samples from Patients Undergoing Autologous Bone Marrow or Peripheral Blood Progenitor Cell Transplantation

Abstract The fate of DNA and protein transgenic sequences in products derived from animals fed transgenic crops has recently raised public interest. Sensitive molecular tests targeting the Bt176 genetic construct and the transgenic Cry1Ab protein were developed to determine whether plant sequences, especially transgenic sequences, are present in animal products. A protocol for total DNA extraction and purification from cow whole blood samples was first drawn up and assessed by spiking with known amounts of DNA from Bt176 maize. The limit of detection for transgenic sequences (35S promoter and Bt176-specific junction sequence) was determined by both the polymerase chain reaction–enzyme-linked immunosorbent assay (PCR–ELISA) and the 5′-nuclease PCR assay. Four additional PCR systems were built to substantiate the results. The first detects a mono-copy maize-specific sequence (ADH promoter). Two others target multi-copy sequences from plant nucleus (26S rRNA gene) and chloroplast (psaB gene). The last one, used as a positive control, targets a mono-copy animal sequence (αs1-casein gene). Both methods detected a minimum spiking at 25 copies of Bt176 maize/mL in 10 mL whole blood samples. The sandwich ELISA kit used detected down to 1 ng transgenic Cry1Ab protein/mL spiked whole blood.

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Comparison between LightCycler Real-Time Polymerase Chain Reaction (PCR) Assay with Serum and PCR-Enzyme-Linked Immunosorbent Assay with Whole Blood Samples for the Diagnosis of Human Brucellosis

Clinical Infectious Diseases ◽

10.1086/426818 ◽

2005 ◽

Vol 40 (2) ◽

pp. 260-264 ◽

Cited By ~ 54

Author(s):

M. I. Queipo-Ortuno ◽

J. D. Colmenero ◽

G. Baeza ◽

P. Morata

Keyword(s):

Polymerase Chain Reaction ◽

Real Time ◽

Whole Blood ◽

Enzyme Linked Immunosorbent Assay ◽

Human Brucellosis ◽

Chain Reaction ◽

Pcr Assay ◽

Blood Samples ◽

Polymerase Chain ◽

Whole Blood Samples

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Screening for Aspergillus spp. using polymerase chain reaction of whole blood samples from patients with haematological malignancies

British Journal of Haematology ◽

10.1046/j.1365-2141.2001.02744.x ◽

2001 ◽

Vol 113 (1) ◽

pp. 180-184 ◽

Cited By ~ 71

Author(s):

Cornelia Lass-Flörl ◽

Johannes Aigner ◽

Eberhard Gunsilius ◽

Andreas Petzer ◽

David Nachbaur ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Whole Blood ◽

Haematological Malignancies ◽

Chain Reaction ◽

Blood Samples ◽

Polymerase Chain ◽

Whole Blood Samples

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Eradication of polymerase chain reaction detectable immunoglobulin gene rearrangement in non-Hodgkin's lymphoma is associated with decreased relapse after autologous bone marrow transplantation

Blood ◽

10.1182/blood.v88.9.3314.bloodjournal8893314 ◽

1996 ◽

Vol 88 (9) ◽

pp. 3314-3322 ◽

Cited By ~ 61

Author(s):

CS Zwicky ◽

AB Maddocks ◽

N Andersen ◽

JG Gribben

Keyword(s):

Bone Marrow ◽

Polymerase Chain Reaction ◽

Bone Marrow Transplantation ◽

Clinical Utility ◽

Autologous Bone Marrow Transplantation ◽

Autologous Bone Marrow ◽

Chain Reaction ◽

Lymphoma Cells ◽

Polymerase Chain ◽

Autologous Bone

In B-cell non-Hodgkin's lymphoma (NHL), as in other B-cell malignancies, clonal rearrangement of the third complementarity determining region (CDR III) of the immunoglobulin heavy chain gene (IgH) provides a useful marker for the detection of minimal residual disease (MRD) after treatment. To determine the clinical utility of IgH polymerase chain reaction (PCR), we analyzed peripheral blood (PB) and bone marrow (BM) samples from 25 patients with NHL with no PCR detectable chromosomal rearrangement who have undergone autologous bone marrow transplantation (ABMT). Patients with histologic bone marrow infiltration at the time of bone marrow harvest were selected for study since this provided us with diagnostic tissue samples. As an initial strategy DNA was amplified using consensus variable (VH) and joining (JH) region primers. In those cases failing to amplify using consensus region primers, PCR was performed using a panel of VH family-specific framework region 1 (FR1) primers. The clonal products were directly sequenced. From the V-N-D region nucleotide sequences, clone specific probes were constructed and used for subsequent detection of MRD. A clonal PCR product could be PCR amplified and directly sequenced in 18 (72%, 90% confidence intervals 54%-86%) of these 25 patients, 8 with diffuse and 10 with follicular NHL. Eight of these 18 patients have relapsed after ABMT. All had detectable lymphoma cells before relapse and the sequence of the CDR III region at the time of relapse was identical to that obtained at the time of ABMT. All 10 patients who remain in complete remission from 18 to 36 months after ABMT had eradication of PCR detectable lymphoma cells after ABMT, although in three patients PCR detectable MRD was detected early after ABMT. We conclude that sequencing and the use of patient specific IgH CDR III oligonucleotides probes provides a simple and highly reliable method to determine the specificity of the IgH PCR technique. The clinical utility of this technique is demonstrated by the finding that eradication of PCR detectable lymphoma cells in these patients is associated with decreased relapse after ABMT (P = .0002).

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Direct Amplification of B1 gene of Toxoplasma gondii DNA using Nested Polymerase Chain Reaction Following Microwave Treatment for Whole Blood Samples

The Iraqi Journal of Veterinary Medicine ◽

10.30539/iraqijvm.v39i1.244 ◽

2015 ◽

Vol 39 (1) ◽

pp. 23-27

Author(s):

Balkes Fadel Hade

Keyword(s):

Polymerase Chain Reaction ◽

Whole Blood ◽

Microwave Treatment ◽

Chain Reaction ◽

Nested Polymerase Chain Reaction ◽

Blood Samples ◽

B1 Gene ◽

Polymerase Chain ◽

Whole Blood Samples ◽

Commercial Kit

Infection with Toxoplasma gondii is a cause of fetal death since T. gondii can be transmitted to the fetus through the placenta (transplacental) from an infected mother or at vaginal delivery. Blood obtained from women and sheep to confirm their infection with toxoplasmosis by using Enzyme Linked Immunosorbant Assay test (ELISA) to ditective positive specific anti-Toxoplasma (IgM, IgG and IgM, or IgG) antibodies. This study used two methodes to extract DNA (the first one was a standard extraction commercial method (CM-PCR) of genomic DNA using a commercial kit (Promega, USA), and the second one was the direct heat DNA extraction using microwave oven (MW-PCR) for whole blood samples obtained from infected women and sheep. Then nested Polymerase Chain Reaction (n PCR) were used to amplify Toxoplasma B1 gene to detect T. gondii DNA in whole blood samples. The results indecated using of microwave treatment instead of commercial kit to extract DNA is low cost and short time,and complement serology for clinical studies and diagnostic purposes of toxoplasmosis.

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RH genotypes among Malaysian blood donors

Asian Biomedicine ◽

10.5372/1905-7415.0804.319 ◽

2014 ◽

Vol 8 (4) ◽

pp. 499-504 ◽

Cited By ~ 1

Author(s):

Rozi Hanisa Musa ◽

Afifah Hassan ◽

Yasmin Ayob ◽

Narazah Mohd Yusoff

Keyword(s):

Polymerase Chain Reaction ◽

Ethnic Groups ◽

Whole Blood ◽

Molecular Basis ◽

Blood Donors ◽

Limited Information ◽

Chain Reaction ◽

Blood Samples ◽

Polymerase Chain ◽

Whole Blood Samples

AbstractBackground: RH genotyping studies have been conducted mainly in people of Caucasian and African descent. There is limited information regarding the molecular basis for RH genotypes in Malaysia.Objectives: To investigate the prevalence and characteristics of RHCE genotypes among different ethnic groups in Malaysia.Methods: A total of 1014 whole blood samples were obtained from donors from 4 different ethnic groups (360 Malays, 434 Chinese, 164 Indians, and 56 others). All samples were phenotyped for C, c, D, E, and e using standard serologic methods and genotyped using polymerase chain reaction (PCR)-based analysis.Results: In the blood samples that we analyzed, the distribution of RH genotype antigens was significantly different among the various ethnic groups. Our findings showed that CCDee is the most common in Malaysian blood donors; 18.4% (187/1014) compared with other genotypes. The ccDEE genotype is more prevalent in the Chinese: 65.6% (82/125), and the ccee genotype is more prevalent in Indians: 47.1% (65/138). There were discrepancies between phenotypes and genotypes. There were 17 (1.7%) discrepancies in RH C/c genotyping results and of these 47% (8/17) occurred in Malays. Discrepancies in RH E/e results occurred in 3 samples (0.3%).Conclusions: Our study provides a database for the distribution of RH genotypes of donors from the major ethnic groups in Malaysia. Methods used in this study are useful for comparing the phenotypes and genotypes. Further investigation should be conducted to study the causes of these discrepancies using other molecular based techniques.

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Detection of Subclinical Systemic Disease in Primary CNS Lymphoma by Polymerase Chain Reaction of the Rearranged Immunoglobulin Heavy-Chain Genes

Journal of Clinical Oncology ◽

10.1200/jco.2006.06.7165 ◽

2006 ◽

Vol 24 (29) ◽

pp. 4754-4757 ◽

Cited By ~ 55

Author(s):

Kristoph Jahnke ◽

Michael Hummel ◽

Agnieszka Korfel ◽

Thomas Burmeister ◽

Philipp Kiewe ◽

...

Keyword(s):

Bone Marrow ◽

Polymerase Chain Reaction ◽

Systemic Disease ◽

Primary Cns Lymphoma ◽

Cns Lymphoma ◽

Chain Reaction ◽

Blood Samples ◽

Pcr Products ◽

Polymerase Chain

Purpose To search for subclinical systemic disease in bone marrow and peripheral blood in patients with primary CNS lymphoma (PCNSL) to elucidate whether extracerebral relapse may represent a sequel of initial occult systemic disease rather than true extracerebral spread. Patients and Methods Bone marrow and peripheral-blood specimens of 24 PCNSL patients were examined using polymerase chain reaction (PCR) for analysis of clonally rearranged immunoglobulin heavy-chain (IgH) genes. Results Identical dominant PCR products were found in bone marrow aspirates, blood samples, and tumor biopsy specimens of two patients, indicating that the same tumor cell population is present in the CNS and in extracerebral sites. Follow-up IgH PCR performed in one of these patients in complete remission 24 months after diagnosis yielded a persistent monoclonal product in the blood. An oligoclonal IgH rearrangement pattern was found in the tumor specimen of two other patients, whereas bone marrow and blood samples demonstrated the same dominant PCR products. Follow-up PCR showed a persistent monoclonal amplificate in blood in one of these patients 27 months after diagnosis. Conclusion It could be demonstrated for the first time that subclinical systemic disease can be present in PCNSL patients at initial diagnosis. Our findings may have an impact on the understanding of PCNSL pathogenesis and the extent of staging and treatment.

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