Quantitation of cytomegalovirus DNA by the polymerase chain reaction as a predictor of disease in solid organ transplantation

2004 ◽  
Vol 73 (2) ◽  
pp. 223-229 ◽  
Author(s):  
Valeria Ghisetti ◽  
Anna Barbui ◽  
Alessandro Franchello ◽  
Silvia Varetto ◽  
Fabrizia Pittaluga ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Organ Transplantation ◽  
Solid Organ Transplantation ◽  
Solid Organ ◽  
Chain Reaction ◽  
Polymerase Chain

Nested Polymerase Chain Reaction With Sequence-Specific Primers Typing for HLA-A, -B, and -C Alleles: Detection of Microchimerism in DR-Matched Individuals

Blood ◽  
1999 ◽  
Vol 94 (4) ◽  
pp. 1471-1477 ◽  
Author(s):  
Anthony S. Carter ◽  
Lucia Cerundolo ◽  
Mike Bunce ◽  
Dicken D.H. Koo ◽  
Kenneth I. Welsh ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Nested Pcr ◽  
Solid Organ Transplantation ◽  
Class Ii ◽  
Hla Class Ii ◽  
Solid Organ ◽  
Chain Reaction ◽  
Specific Primers ◽  
Polymerase Chain ◽  
Hla Class Ii Alleles

Abstract It is widely accepted that donor leukocytes survive within the recipient periphery after blood transfusion or solid organ transplantation. The significance of this microchimerism remains unclear, partially because of the insecurity of assays used to detect the donor-derived material. The techniques used to detect donor-derived DNA within recipient peripheral blood rely largely on major histocompatibility complex class II polymorphism. We and others have shown that the sensitivity of polymerase chain reaction with sequence-specific primers (PCR-SSP) typing for HLA class II alleles can be increased 100-fold by the addition of a primary amplification step (nested PCR-SSP). We have now extended this technique to encompass typing for HLA class I alleles, thereby adding flexibility to microchimerism testing by enabling testing of recipients HLA-DR matched with their donors. However, the high level of sensitivity achieved with the technique (1:100,000) leads to a concomitant decrease in the specificity that results in the amplification of unexpected products, a phenomenon we encountered in the development of our nested PCR-SSP typing system for HLA class II alleles. We describe here how it is possible to compensate for these anomalies by including multiple testing of a pretransfusion sample that acts as a specificity control, establishing a rigorous baseline for subsequent analysis.

Nested Polymerase Chain Reaction With Sequence-Specific Primers Typing for HLA-A, -B, and -C Alleles: Detection of Microchimerism in DR-Matched Individuals

Blood ◽  
1999 ◽  
Vol 94 (4) ◽  
pp. 1471-1477
Author(s):  
Anthony S. Carter ◽  
Lucia Cerundolo ◽  
Mike Bunce ◽  
Dicken D.H. Koo ◽  
Kenneth I. Welsh ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Nested Pcr ◽  
Solid Organ Transplantation ◽  
Class Ii ◽  
Hla Class Ii ◽  
Solid Organ ◽  
Chain Reaction ◽  
Specific Primers ◽  
Polymerase Chain ◽  
Hla Class Ii Alleles

It is widely accepted that donor leukocytes survive within the recipient periphery after blood transfusion or solid organ transplantation. The significance of this microchimerism remains unclear, partially because of the insecurity of assays used to detect the donor-derived material. The techniques used to detect donor-derived DNA within recipient peripheral blood rely largely on major histocompatibility complex class II polymorphism. We and others have shown that the sensitivity of polymerase chain reaction with sequence-specific primers (PCR-SSP) typing for HLA class II alleles can be increased 100-fold by the addition of a primary amplification step (nested PCR-SSP). We have now extended this technique to encompass typing for HLA class I alleles, thereby adding flexibility to microchimerism testing by enabling testing of recipients HLA-DR matched with their donors. However, the high level of sensitivity achieved with the technique (1:100,000) leads to a concomitant decrease in the specificity that results in the amplification of unexpected products, a phenomenon we encountered in the development of our nested PCR-SSP typing system for HLA class II alleles. We describe here how it is possible to compensate for these anomalies by including multiple testing of a pretransfusion sample that acts as a specificity control, establishing a rigorous baseline for subsequent analysis.

scholarly journals Outcome of COVID-19 in Solid Organ Malignancies: Experience From a Tertiary Cancer Center in Eastern India

2021 ◽  
pp. 1374-1379
Author(s):  
Somnath Roy ◽  
Joydeep Ghosh ◽  
Sandip Ganguly ◽  
Debapriya Mondal ◽  
Deepak Dabkara ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcriptase ◽  
Median Duration ◽  
Cancer Center ◽  
Nasopharyngeal Swab ◽  
Solid Organ ◽  
Chain Reaction ◽  
Unique Challenge ◽  
Risk Of Death ◽  
Polymerase Chain

PURPOSE The COVID-19 pandemic has imposed a unique challenge to oncology patients. Outcome data on COVID-19 in patients with cancer from the Indian subcontinent are scarce in the literature. We aimed to evaluate the outcome of patients with COVID-19 on active systemic anticancer therapy. MATERIALS AND METHODS This is a retrospective study of patients with solid organ malignancies undergoing systemic therapy with a diagnosis of COVID-19 between March 2020 and February 2021. COVID-19 was diagnosed if a reverse transcriptase polymerase chain reaction assay from oropharyngeal or nasopharyngeal swab was positive for severe acute respiratory syndrome coronavirus 2. The objectives were to evaluate the outcome of COVID-19 and factors predicting the outcome. RESULTS A total of 145 patients were included with a median age of 58 years (range, 20-81 years). Treatment was curative in 60 (42%) patients. Of all symptomatic cases (n = 88, 61%), 50 had mild, 27 had moderate and 19 had severe COVID-19–related symptoms as per WHO criteria. Fifty (34%) patients required hospitalization with a median duration of hospital stay of 12 days (range, 4-25 days); five patients required intensive care unit admission. The rest were treated with home isolation and did not require further hospitalization. Twenty-two (15%) patients died, and the risk of death was significantly associated with severity of symptoms (odds ratio, 91.3; 95% CI, 9.1 to 919.5, P = .0001) but not with any other clinical factors. Drug holiday was given to 63 (44%) patients with a median duration of 25 days (range, 7-88 days). The median duration to reverse transcriptase polymerase chain reaction–negative was 16 days (range, 7-62 days). CONCLUSION COVD-19–related death rate was 15% among patients with solid organ malignancies. The severity of the symptoms was related to mortality. The majority of patients with mild symptoms were treated at home isolation.

scholarly journals Monitoring of cytomegalovirus (CMV) infection in solid organ transplant recipients: quantitation of CMV DNAemia by two real-time polymerase chain reaction assays

2016 ◽  
Vol 31 (3) ◽  
Author(s):  
Angela Chiereghin ◽  
Giulia Piccirilli ◽  
Gabriele Turello ◽  
Diego Squarzoni ◽  
Claudia Pavia ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Result ◽  
Lower Amount ◽  
Solid Organ ◽  
Transplant Recipients ◽  
Chain Reaction ◽  
Clinical Value ◽  
Polymerase Chain ◽  
Cmv Dnaemia

<em>Background and aim:</em> Quantification of cytomegalovirus (CMV) DNAemia is essential in clinical management of post-transplant infection. We evaluated the performances of two quantitative real-time polymerase chain reaction (PCR) assays. <br /><em>Materials and Methods</em>: 114 serial whole blood samples collected from 14 actively infected transplant recipients were processed by Abbott RealTime CMV PCR kit (Abbott Molecular) and CMV ELITe MGB™ kit (ELITech Group). The Quality Control for Molecular Diagnostics human CMV panels was also tested. <br /><em>Results</em>: Sixteen (14%) samples resulted negative and 59 (51.7%) positive with a quantitative result for both assays. In the 59 samples, the coefficient of correlation was 0.856. Bland-Altman analysis showed a mean difference of &lt;0.11 log10 copies/mL (standard deviation=0.38 log10 copies/mL). The assays gave CMV-DNA loads differing by 1 log10 DNA copies/mL in 57 samples (96.6%) and by &lt;0.5 log10 DNA copies/mL in 48 samples (81.3%). Eleven (9.6%) samples were positive with a quantitative result with Abbott and negative with ELITech. Sixteen (14%) positive samples with a quantitative result for Abbott resulted positive but below the lower limit of quantification (LLQ) for ELITech. Twelve (10.5%) samples resulted negative with ELITech and positive but below the LLQ with Abbott. No samples were positive with ELITech and negative with Abbott. <br /><em>Conclusions</em>: The assays showed a good correlation between CMVDNA levels detected and variation in CMV-DNA &lt;0.5 log10 was observed in the majority of the samples. The viral load kinetic profiles of the assays were overlapping in all patients, but Abbott showed higher sensitivity in samples containing lower amount of DNA. The clinical value of this greater sensitivity requires further investigation.

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