Effect of heparin on polymerase chain reaction

Satsangi, Jack; Jewell, D P; Welsh, Ken I.; Bunce, Michael; Bell, John I.

doi:10.1016/s0140-6736(94)92622-0

Cited by 121 publications

(81 citation statements)

References 4 publications

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“…It is also possible that PCR amplification of ASFV DNA from blood was reduced or blocked by the presence of PCR-inhibitory substances. PCR inhibitors in blood have been identified and include natural blood components, mainly heme (3) and leukocyte DNA (21), or added anticoagulants, such as heparin (28).…”

Section: Resultsmentioning

confidence: 99%

Preclinical Diagnosis of African Swine Fever in Contact-Exposed Swine by a Real-Time PCR Assay

et al. 2005

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A fluorogenic probe hydrolysis (TaqMan) PCR assay for African swine fever virus (ASFV) was developed and evaluated in experimentally infected swine. This sensitive and specific one-step single-tube assay, which can be performed in 2 h or less, detected viral DNA in tonsil scraping samples 2 to 4 days prior to onset of clinical disease. Thus, the assay would have application for preclinical diagnosis of African swine fever and surveillance and/or emergency management of a disease outbreak

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Section: Resultsmentioning

confidence: 99%

Preclinical Diagnosis of African Swine Fever in Contact-Exposed Swine by a Real-Time PCR Assay

et al. 2005

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“…Not all samples could be HLA typed due to a large number of assay failures secondary to heparinisation, which can inhibit HLA typing [6]. We typed 200 sporadic, 126 LDNN and 182 nephropathy patients, using the phototyping method [7] which involves typing for HLA-A, B, C, DR, DQA1 and DQB1 using a mix of 144 sequence specific primers.…”

Section: Methodsmentioning

confidence: 99%

Human leucocyte antigen and insulin gene regions and nephropathy in Type I diabetes

et al. 1999

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The human leucocyte antigen (HLA) region has a major influence on susceptibility to Type I diabetes [1] and is possibly implicated in susceptibility to diabetic complications. Human leucocyte antigen-identical, non-diabetic siblings of diabetic probands have glomerular basement membrane expansion, which is not observed in HLA-non-identical siblings [2]. This is also observed in DR4 positive parents of diabetic patients, suggesting that even in the absence of diabetes, microangiopathy may occur due to HLA-related factors [2]. Previous examination of HLA in nephropathy has shown positive and negative associations with A2, B8, B15, DR4 and DR3/4 [3]. The insulin gene (INS) is also associated with Type I diabetes, and has been implicated in the onset of vascular disease [4]. As macrovascular disease is common in nephropathy, INS is a suitable candidate gene for susceptibility to nephropathy. Previous studies suggest an excess of class I allele homozygotes in patients with nephropathy [4].Studies examining the role of HLA or INS have often been small and ascertainment of nephropathy status has been poor. The aim of our study was to clarify the role of these genes in susceptibility to nephropathy in three large, well characterised cohorts with Type I diabetes. Diabetologia (1999) Abstract Aims/hypothesis. Diabetic nephropathy seems to have a strong genetic component. Genes involved in the genetic susceptibility to Type I (insulin-dependent) diabetes have been suggested to have a role in the development of diabetic nephropathy. This study aimed to examine the role of human leucocyte antigen and insulin genes in susceptibility to nephropathy in patients with Type I diabetes. Methods. We carried out a genetic association study examining insulin gene polymorphisms using three large cohorts of patients with Type I diabetes: nephropathy (n = 258), long duration non-nephropathy (n = 153) and a recently diagnosed (sporadic) diabetic cohort (n = 264). Human leucocyte antigen typing results were obtained in a smaller number due to assay failures (n = 182, 126 and 200 respectively).

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“…8,9 Among the most potent PCR inhibitors reported are hemoglobin/heme, 10 leukocyte DNA, 11 and an IgG fraction, 12 in addition to anticoagulants such as EDTA, sodium citrate, and heparin, which also inhibit PCR. 13 Other known inhibitors are bilirubin, bile salts, and lactoferrin.…”

mentioning

confidence: 99%

Direct DNA Amplification from Crude Clinical Samples Using a PCR Enhancer Cocktail and Novel Mutants of Taq

Zhang

Kermekchiev

Barnes

2010

The Journal of Molecular Diagnostics

124

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PCR-based clinical and forensic tests often have low sensitivity or even false-negative results caused by potent PCR inhibitors found in blood and soil. It is widely accepted that purification of target DNA before PCR is necessary for successful amplification. In an attempt to overcome PCR inhibition , enhance PCR amplification , and simplify the PCR protocol, we demonstrate improved PCR-enhancing cocktails containing nonionic detergent , L-carnitine , D-(؉)-trehalose , and heparin. These cocktails , in combination with two inhibitor-resistant Taq mutants , OmniTaq and Omni Klentaq , enabled efficient amplification of exogenous , endogenous , and high-GC content DNA targets directly from crude samples containing human plasma , serum , and whole blood without DNA purification. In the presence of these enhancer cocktails , the mutant enzymes were able to tolerate at least 25% plasma , serum , or whole blood and as high as 80% GC content templates in PCR reactions. These enhancer cocktails also improved the performance of the novel Taq mutants in real-time PCR amplification using crude samples , both in SYBR Green fluorescence detection and TaqMan assays. The novel enhancer mixes also facilitated DNA amplification from crude samples with various commercial

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Effect of heparin on polymerase chain reaction

Cited by 121 publications

References 4 publications

Preclinical Diagnosis of African Swine Fever in Contact-Exposed Swine by a Real-Time PCR Assay

Preclinical Diagnosis of African Swine Fever in Contact-Exposed Swine by a Real-Time PCR Assay

Human leucocyte antigen and insulin gene regions and nephropathy in Type I diabetes

Direct DNA Amplification from Crude Clinical Samples Using a PCR Enhancer Cocktail and Novel Mutants of Taq

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