Comparison of five protocols to extract DNA from paraffin-embedded tissues for the detection of human papillomavirus

Aldana, Adalucy Álvarez; Martínez, José William; Sepúlveda-Arias, Juan Carlos

doi:10.1016/j.prp.2014.10.011

Cited by 23 publications

(30 citation statements)

References 53 publications

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“…It is likely that the anomaly is a likely consequence of one of the reasons why ∆Cq and DIN do not correlate as discussed above. Different kits have various efficiencies at removing inhibitors when purifying DNA from FFPE sections [23][24][25] , so the ExScale system might simply be less efficient at removing inhibitors than the QIAamp kit. We could not perform 260:280 and 260:230 purity analyses on ExScale-purified DNA because ExScale do not provide sufficient DNA elution buffer to blank the spectrophotometer (a significant disadvantage of ExScale in our view).…”

Section: Discussionmentioning

confidence: 99%

Extracting DNA from FFPE Tissue Biospecimens Using User-Friendly Automated Technology: Is There an Impact on Yield or Quality?

Mathieson

Guljar

Sánchez

et al. 2018

Biopreservation and Biobanking

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DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue blocks is amenable to analytical techniques, including sequencing. DNA extraction protocols are typically long and complex, often involving an overnight proteinase K digest. Automated platforms that shorten and simplify the process are therefore an attractive proposition for users wanting a faster turn-around or to process large numbers of biospecimens. It is, however, unclear whether automated extraction systems return poorer DNA yields or quality than manual extractions performed by experienced technicians. We extracted DNA from 42 FFPE clinical tissue biospecimens using the QiaCube (Qiagen) and ExScale (ExScale Biospecimen Solutions) automated platforms, comparing DNA yields and integrities with those from manual extractions. The QIAamp DNA FFPE Spin Column Kit was used for manual and QiaCube DNA extractions and the ExScale extractions were performed using two of the manufacturer's magnetic bead kits: one extracting DNA only and the other simultaneously extracting DNA and RNA. In all automated extraction methods, DNA yields and integrities (assayed using DNA Integrity Numbers from a 4200 TapeStation and the qPCR-based Illumina FFPE QC Assay) were poorer than in the manual method, with the QiaCube system performing better than the ExScale system. However, ExScale was fastest, offered the highest reproducibility when extracting DNA only, and required the least intervention or technician experience. Thus, the extraction methods have different strengths and weaknesses, would appeal to different users with different requirements, and therefore, we cannot recommend one method over another.

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Section: Discussionmentioning

confidence: 99%

Extracting DNA from FFPE Tissue Biospecimens Using User-Friendly Automated Technology: Is There an Impact on Yield or Quality?

Mathieson

Guljar

Sánchez

et al. 2018

Biopreservation and Biobanking

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“…To monitor the quality of DNA extraction and to verify the presence of amplifiable DNA in our study, an endogenous DNA control gene HBB was quantified in FFPE and FF tissue (Fig1) [ 23 , 42 , 43 ]. Not surprisingly, the highest amount of HBB was found in the FF tissue, with Ct values of about 25 which indicates high amounts of high quality DNA as starting point for PCR amplification.…”

Section: Discussionmentioning

confidence: 99%

Traceability and distribution of Neisseria meningitidis DNA in archived post mortem tissue samples from patients with systemic meningococcal disease

et al. 2017

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BackgroundThe pathophysiology and outcome of meningococcal septic shock is closely associated with the plasma level of N. meningitidis lipopolysaccharides (LPS, endotoxin) and the circulating level of meningococcal DNA. The aim of the present study was to quantify the number of N. meningitidis in different formalin-fixed, paraffin-embedded (FFPE) tissue samples and fresh frozen (FF) tissue samples from patients with systemic meningococcal disease (SMD), to explore the distribution of N. meningitidis in the body.MethodsDNA in FFPE and FF tissue samples from heart, lungs, liver, kidneys, spleen and brain from patients with meningococcal shock and controls (lethal pneumococcal infection) stored at variable times, were isolated. The bacterial load of N. meningitidis DNA was analyzed using quantitative real-time PCR (qPCR) and primers for the capsule transport A (ctrA) gene (1 copy per N. meningitidis DNA). The human beta-hemoglobin (HBB) gene was quantified to evaluate effect of the storage times (2-28 years) and storage method in archived tissue.Results N. meningitidis DNA was detected in FFPE and FF tissue samples from heart, lung, liver, kidney, and spleen in all patients with severe shock. In FFPE brain, N. meningitidis DNA was only detected in the patient with the highest concentration of LPS in the blood at admission to hospital. The highest levels of N. meningitidis DNA were found in heart tissue (median value 3.6 × 107 copies N. meningitidis DNA/μg human DNA) and lung tissue (median value 3.1 × 107 copies N. meningitidis DNA/μg human DNA) in all five patients. N. meningitidis DNA was not detectable in any of the tissue samples from two patients with clinical meningitis and the controls (pneumococcal infection). The quantity of HBB declined over time in FFPE tissue stored at room temperature, suggesting degradation of DNA.ConclusionsHigh levels of N. meningitidis DNA were detected in the different tissue samples from meningococcal shock patients, particularly in the heart and lungs suggesting seeding and major proliferation of meningococci in these organs during the development of shock, probably contributing to the multiple organ failure. The age of archived tissue samples appear to have an impact on the amount of quantifiable N. meningitidis DNA.

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“…In addition, 60.7% of study specimens were HNC from oral cavity or larynx, which are driven more by alcohol and smoking than HPV [4]. Furthermore, HPV prevalence rate in cervical cancer biopsies appear lower than that reported in other studies [16], suggesting possible low detection sensitivity due to the storage protocol of biopsy samples into paraffin, as mentioned elsewhere [17].…”

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confidence: 99%

Infrequent detection of human papillomavirus infection in head and neck cancers in the Central African Republic: a retrospective study

Kofi¹,

Mossoro-Kpinde²,

Bouassa³

et al. 2019

Infect Agents Cancer

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We carried out a retrospective study on the prevalence of HPV and genotype distribution by nested PCR and nucleotide sequencing analysis, in formalin-fixed, paraffin-embedded biopsies of 135 head and neck cancers (HNC) and 29 cervical cancers received between 2009 and 2017 for diagnosis at the Laboratoire National de Biologie Clinique et de Santé Publique of Bangui, the capital city of the Central African Republic. One oropharyngeal squamous cell carcinoma sample was positive for HPV type 16. The overall HPV prevalence in HNC biopsies was 0.74% (95% CI: 0.0–2.2). Among the 29 cervical cancer samples, 19 (65.5%; 95% CI: 48.2–82.8) were positive for HPV. These results indicate that HNC are infrequently associated with HPV infection in the Central African Republic.

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Comparison of five protocols to extract DNA from paraffin-embedded tissues for the detection of human papillomavirus

Cited by 23 publications

References 53 publications

Extracting DNA from FFPE Tissue Biospecimens Using User-Friendly Automated Technology: Is There an Impact on Yield or Quality?

Extracting DNA from FFPE Tissue Biospecimens Using User-Friendly Automated Technology: Is There an Impact on Yield or Quality?

Traceability and distribution of Neisseria meningitidis DNA in archived post mortem tissue samples from patients with systemic meningococcal disease

Infrequent detection of human papillomavirus infection in head and neck cancers in the Central African Republic: a retrospective study

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