RESUMO
BACKGROUND: Head and neck squamous cell carcinomas are treated by surgery, radiotherapy (RT), chemoradiotherapy (CRT) or combinations thereof, but locoregional recurrences (LRs) occur in 30-40% of treated patients. We have previously shown that in approximately half of the LRs after CRT, cancer driver mutations are not shared with the index tumor. AIM: To investigate two possible explanations for these genetically unrelated relapses, treatment-induced genetic changes and intratumor genetic heterogeneity. METHODS: To investigate treatment-induced clonal DNA changes, we compared copy number alterations (CNAs) and mutations between primary and recurrent xenografted tumors after treatment with (C)RT. Intratumor genetic heterogeneity was studied by multi-region sequencing on DNA from 31 biopsies of 11 surgically treated tumors. RESULTS: Induction of clonal DNA changes by (C)RT was not observed in the xenograft models. Multi-region sequencing demonstrated variations in CNA profiles between paired biopsies of individual tumors, with copy number heterogeneity scores varying from 0.027 to 0.333. In total, 32 cancer driver mutations could be identified and were shared in all biopsies of each tumor. Remarkably, multi-clonal mutations in these same cancer driver genes were observed in 6 of 11 tumors. Genetically distinct heterogeneous cell cultures could also be established from single tumors, with different biomarker profiles and drug sensitivities. CONCLUSION: Intratumor genetic heterogeneity at the level of the cancer driver mutations might explain the discordant mutational profiles in LRs after CRT, while there are no indications in xenograft models that these changes are induced by CRT.
Assuntos
Heterogeneidade Genética , Neoplasias de Cabeça e Pescoço , Humanos , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/terapia , Mutação , Recidiva , DNARESUMO
Organ and tissue donation depends on non-transplant clinicians to identify and timeously refer potential donors and to counsel families compassionately about the prognosis at end of life. Organ donation referral is often felt to be beyond the capacity of district-level hospital services. In this case series, we report on four referrals from a geographically remote, public sector district-level hospital, and review the identification, referral and consent process of potential donors after brain death, and also donors after circulatory death. For the one successfully consented donor we report on the donor work-up and management, and the outcome of the organ recovery and organ allocation process.
Assuntos
Transplante de Órgãos , Obtenção de Tecidos e Órgãos , Humanos , África do Sul , Doadores de Tecidos , DocentesRESUMO
A previously healthy 10-year-old girl, living in a sheep-farming community in South Africa with exposure to dogs, presented to her local hospital with generalised tonic-clonic seizures. The initial clinical assessment and laboratory work-up were unremarkable. When she presented with further seizures 6 months later, attempts to arrange neuroimaging and specialist assessment were unsuccessful owing to restrictions on routine healthcare services during the SARS-CoV-2 nationwide lockdown. Subsequently, 11 months after her first presentation, she developed focal neurological signs suggestive of raised intracranial pressure. A brain computed tomography scan revealed a left-sided cerebral cyst and imminent tonsillar herniation. An emergency burr-hole procedure was performed to relieve the raised intracranial pressure, followed by definitive neurosurgical excision of cysts. Hydatid protoscolices and hooklets were seen on microscopy of cyst fluid, and treatment with albendazole and praziquantel was initiated. While her infection was treated successfully, long-term sequelae including permanent blindness and hemiparesis could potentially have been prevented with early neuroimaging and surgical intervention.
Assuntos
Anticestoides/administração & dosagem , Encefalopatias/diagnóstico , COVID-19 , Equinococose/diagnóstico , Albendazol/administração & dosagem , Encefalopatias/tratamento farmacológico , Encefalopatias/parasitologia , Criança , Diagnóstico Tardio , Equinococose/tratamento farmacológico , Feminino , Humanos , Hipertensão Intracraniana/parasitologia , Praziquantel/administração & dosagem , Convulsões/parasitologia , África do Sul , Tomografia Computadorizada por Raios XRESUMO
AIMS: Diagnostic and post-induction 123I-meta-iodobenzylguanidine (123I-mIBG) scans have prognostic significance in the treatment of neuroblastoma, but data from low- and middle-income countries are limited due to resource constraints. The aim of this study was to determine the association between neuroblastoma-associated tumour markers (lactate dehydrogenase [LDH], ferritin and MYCN amplification) and 123I-mIBG scans (modified Curie scores and metastatic disease patterns) in predicting complete metastatic response rates (mCR) and overall survival. MATERIALS AND METHODS: Two hundred and ninety patients diagnosed with high-risk neuroblastoma in South Africa between January 2000 and May 2018 and a subanalysis of 78 patients with diagnostic 123I-mIBG scans were included. Data collection included LDH, ferritin and MYCN amplification at diagnosis. Two nuclear physicians independently determined the modified Curie scores and pattern of distribution for each diagnostic and post-induction 123I-mIBG scans with high inter-rater agreement (r = 0.952) and reliability (K = 0.805). The cut-off values for the diagnostic and post-induction modified Curie scores of ≥7.0 (P = 0.026) and 3 (P = 0.009), respectively, were generated. The association between the tumour markers and the modified Curie score of the 123I-mIBG scans was determined using post-induction mCR and 2-year overall survival. RESULTS: Diagnostic LDH (P < 0.001), ferritin (P < 0.001) and the diagnostic modified Curie scores (P = 0.019) significantly predicted mCR. Only ferritin correlated with diagnostic modified Curie scores (P = 0.003) but had a low correlation coefficient of 0.353. On multivariable analysis, the only significant covariate for 2-year overall survival at diagnosis was LDH <750 U/l (P = 0.024). A post-induction chemotherapy modified Curie score ≤3.0 had a 2-year overall survival of 46.2% compared with 30.8% for a score >3.0 (P = 0.484). CONCLUSION: LDH, ferritin and the diagnostic 123I-mIBG scans significantly predicted mCR, but only LDH predicted 2-year overall survival. Ferritin and the modified Curie scores correlated with each other. MYCN amplification neither correlated with any aspect of the 123I-mIBG scans nor significantly predicted mCR or 2-year overall survival. LDH and ferritin are therefore appropriate neuroblastoma tumour markers to be used in low- and middle-income countries with limited or no access to mIBG scans and/or MYCN amplification studies.
Assuntos
3-Iodobenzilguanidina , Neuroblastoma , Biomarcadores Tumorais/genética , Criança , Humanos , Neuroblastoma/diagnóstico por imagem , Neuroblastoma/genética , Cintilografia , Reprodutibilidade dos TestesRESUMO
[This corrects the article DOI: 10.1186/s13017-016-0089-y.].
RESUMO
Drug-induced liver injury (DILI) is a leading cause of acute hepatic failure and a major reason for market withdrawal of drugs. Idiosyncratic DILI is multifactorial, with unclear dose-dependency and poor predictability since the underlying patient-related susceptibilities are not sufficiently understood. Because of these limitations, a pharmaceutical research option would be to reduce the compound-related risk factors in the drug-discovery process. Here we describe the development and validation of a methodology for the assessment of DILI risk of drug candidates. As a training set, 81 marketed or withdrawn compounds with differing DILI rates - according to the FDA categorization - were tested in a combination of assays covering different mechanisms and endpoints contributing to human DILI. These include the generation of reactive metabolites (CYP3A4 time-dependent inhibition and glutathione adduct formation), inhibition of the human bile salt export pump (BSEP), mitochondrial toxicity and cytotoxicity (fibroblasts and human hepatocytes). Different approaches for dose- and exposure-based calibrations were assessed and the same parameters applied to a test set of 39 different compounds. We achieved a similar performance to the training set with an overall accuracy of 79% correctly predicted, a sensitivity of 76% and a specificity of 82%. This test system may be applied in a prospective manner to reduce the risk of idiosyncratic DILI of drug candidates.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos/métodos , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Animais , Calibragem , Glutationa/metabolismo , Humanos , Camundongos , Células NIH 3T3RESUMO
BACKGROUND: High-risk (hr) human papillomavirus (HPV) infections play a causal role in the development of cervical cancer. The detection of hrHPV is, therefore, advocated in cervical cancer screening programs. OBJECTIVES: The aim of this study was to determine the performance of a novel HPV typing assay, PapillomaFinder® SMART 20. This is a one-tube-per-sample method, to be performed on standard real-time PCR platforms, using melting curve analysis to distinguish targets. The assay detects all 14 hrHPV types, of which 16, 18, 31, 33, 35, 39, 45, 52, 56 and 58 individually. HrHPV types 51, 59, 66 and 68 are detected in an hrHPV pool, and low-risk (lr) HPV types 6, 11, 40, 42, 43 and 44 in an lrHPV pool. STUDY DESIGN: The method was tested on HPV plasmid models, WHO and QCMD proficiency panels and a series of clinical cytological samples (n=45), the latter in comparison with a clinically validated real-time quantitative PCR. RESULTS: Type-specificity of the test was 100% using plasmids, the WHO and QCMD panels. Sensitivity for hrHPV in single infections was 100% using the WHO and QCMD panels and cytological samples, with an analytical sensitivity of 10-25 copies per reaction for all HPV types tested. Of the 34 HPV types present in the 8 multiple infections in the WHO panel, 30 were detected. In all cytological samples at least one hrHPV type was found, in concordance with the clinically validated method. Only when the viral load of the dominant HPV types in multiple infections greatly exceeded that of the other types in the infection, those other types were not always detected. CONCLUSIONS: PapillomaFinder® SMART 20 is a rapid, easy to perform, single tube HPV typing assay. The assay detects the 14 hrHPV types, and the 6 most important lrHPV types with a high sensitivity and type-specificity.
Assuntos
Técnicas de Genotipagem/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Papillomaviridae/classificação , Papillomaviridae/genética , Infecções por Papillomavirus/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Feminino , Humanos , Infecções por Papillomavirus/virologia , Sensibilidade e Especificidade , Fatores de TempoRESUMO
OBJECTIVES: To study the use of limb perfusion scans in children with limb-threatening ischaemia and determine whether such scans are helpful in making clinical decisions. METHODS: This retrospective study compared the clinical, scan and surgical findings in children who had limb perfusion scans for critical limb ischaemia at Red Cross War Memorial Children's Hospital, Cape Town, South Africa, from July 2001 to December 2010. Records were reviewed and the data analysed for aetiology, clinical findings, limb perfusion results, operative findings and outcome. RESULTS: There were complete clinical and scan records for 20/22 patients, aged 1 month to 12 years. The causes of limb ischaemia were meningococcal septicaemia (n = 9), septic shock (n = 6), hypovolaemic shock due to gastroenteritis (n = 4), and electrical burns (n = 1). The clinical, scan and surgical findings correlated in 40/48 imaged limbs. In one leg the findings did not correlate, but the perfusion scan results predicted the outcome. In the remaining seven cases the exact correlation was uncertain owing to technical difficulties or absent operative notes. CONCLUSION: This study describes a method for performing limb perfusion studies in children. Limb perfusion studies correlated well with surgical findings. These studies were useful in treatment decisions, parent and patient counselling and surgical planning. They supplemented clinical examination in assessment of the children.
Assuntos
Isquemia/diagnóstico , Isquemia/etiologia , Extremidade Inferior/irrigação sanguínea , Fatores Etários , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Isquemia/cirurgia , Masculino , Imagem de Perfusão , Valor Preditivo dos Testes , Estudos Retrospectivos , África do Sul , Resultado do TratamentoRESUMO
OBJECTIVE: A systematic review of randomized clinical trials (RCTs) to assess the additional value of hyperbaric oxygen therapy (HBOT) in promoting the healing of diabetic foot ulcers and preventing amputations was performed. METHODS: MEDLINE, Embase, and the Cochrane Library were searched to identify RCTs in patients with diabetic foot ulcers published up to August 2013. Eligible studies reported the effectiveness of adjunctive HBOT with regard to wound healing, amputations, and additional interventions. RESULTS: Seven of the 669 identified articles met the inclusion criteria, comprising 376 patients. Three trials included 182 patients with ischaemic ulcers, two trials studied 64 patients with non-ischaemic ulcers, and two trials comprising 130 patients did not specify ulcer type. Two trials were of good methodological quality. Pooling of data was deemed inappropriate because of heterogeneity. Two RCTs in patients with ischaemic ulcers found increased rates of complete healing at 1-year follow-up (number needed to treat (NNT) 1.8 (95% CI: 1.1 to 4.6) and 4.1 (95% CI: 2.3 to 19)), but found no difference in amputation rates. A third trial in ischaemic ulcers found significantly lower major amputation rates in patients with HBOT (NNT 4.2, 95% CI: 2.4 to 17), but did not report on wound healing. None of the RCTs in non-ischaemic ulcers reported differences in wound healing or amputation rates. Two trials with unknown ulcer types reported beneficial effects on amputation rates, although the largest trial used a different definition for both outcomes. HBOT did not influence the need for additional interventions. CONCLUSION: Current evidence shows some evidence of the effectiveness of HBOT in improving the healing of diabetic leg ulcers in patients with concomitant ischaemia. Larger trials of higher quality are needed before implementation of HBOT in routine clinical practice in patients with diabetic foot ulcers can be justified.
Assuntos
Pé Diabético/terapia , Oxigenoterapia Hiperbárica , Isquemia/terapia , Cicatrização , Amputação Cirúrgica , Terapia Combinada , Pé Diabético/diagnóstico , Pé Diabético/fisiopatologia , Humanos , Isquemia/diagnóstico , Isquemia/fisiopatologia , Salvamento de Membro , Fluxo Sanguíneo Regional , Resultado do TratamentoRESUMO
Our aim was to investigate whether high-risk HPV (hrHPV) mRNA detection by PreTect HPV-Proofer can be used to stratify hrHPV DNA-positive women of different cytology classes for risk of high-grade cervical intraepithelial neoplasia or worse (cervical precancer or cancer, i.e., cervical intraepithelial neoplasia grade 2 or higher [≥ CIN2]). A total of 375 women participating in population-based screening, with a GP5+/6+-PCR hrHPV DNA-positive cervical scrape with normal cytology (n = 202), borderline or mild dyskaryosis (BMD) (n = 88), or moderate dyskaryosis or worse (>BMD) (n = 85), were enrolled. Cervical scrapes were additionally subjected to HPV16/18/31/33/45 E6/E7 mRNA analysis by PreTect HPV-Proofer (mRNA test). Referral and follow-up policies were based on cytology, hrHPV DNA, and mRNA testing. The primary study endpoint was the number of ≥CIN2 detected within 3 years of follow-up. The mRNA positivity increased with the severity of cytological abnormality, ranging from 32% (64/202) in hrHPV DNA-positive women with normal cytology to 47% (41/88) in BMD and 68% (58/85) in >BMD groups (P < 0.01). Women with ≥ CIN2 were more likely to test positive by mRNA test (63%) than women without evidence of ≥ CIN2 (32%; P < 0.01). A positive mRNA test result conferred an increased ≥ CIN2 risk in hrHPV DNA-positive women with normal cytology, i.e., 0.55 (95% confidence interval [95% CI], 0.34 to 0.76) in mRNA-positive versus 0.20 (95% CI, 0.07 to 0.33) in mRNA-negative women. In hrHPV DNA-positive women with BMD or >BMD, the result of the mRNA test did not influence the ≥ CIN2 risk. In conclusion, mRNA testing by PreTect HPV-Proofer might be of value to select hrHPV DNA-positive women with normal cytology in need of immediate referral for colposcopy.
Assuntos
Técnicas de Diagnóstico Molecular/métodos , Proteínas Oncogênicas Virais/biossíntese , Infecções por Papillomavirus/diagnóstico , RNA Mensageiro/análise , RNA Viral/análise , Neoplasias do Colo do Útero/diagnóstico , Virologia/métodos , Adulto , Idoso , Técnicas Citológicas/métodos , Feminino , Humanos , Pessoa de Meia-Idade , Proteínas Oncogênicas Virais/genética , Infecções por Papillomavirus/virologia , RNA Mensageiro/genética , RNA Mensageiro/isolamento & purificação , RNA Viral/genética , RNA Viral/isolamento & purificação , Neoplasias do Colo do Útero/virologiaRESUMO
Tigecycline, the first of a new class of broad-spectrum antibiotics (the glycylcyclines), has been licensed in South Africa for the parenteral treatment of adult patients with complicated intra-abdominal infections (cIAIs) and complicated skin and soft-tissue infections (cSSTIs). This article serves as a summary of the guideline on the appropriate use of tigecycline, published in mid-2010 as a collaborative effort by representatives of the Association of Surgeons of South Africa, the Critical Care Society of Southern Africa, the Federation of Infectious Diseases Societies of Southern Africa, the South African Thoracic Society and the Trauma Society of South Africa.
Assuntos
Antibacterianos/uso terapêutico , Minociclina/análogos & derivados , Guias de Prática Clínica como Assunto , Tratamento Farmacológico/normas , Quimioterapia Combinada/normas , Humanos , Minociclina/uso terapêutico , TigeciclinaAssuntos
Antibacterianos/uso terapêutico , Infecções Bacterianas/tratamento farmacológico , Farmacorresistência Bacteriana , Promoção da Saúde/organização & administração , Medicamentos sob Prescrição/uso terapêutico , Atenção Primária à Saúde/organização & administração , Infecções Bacterianas/epidemiologia , Humanos , Programas Nacionais de Saúde/organização & administração , África do SulAssuntos
Controle de Doenças Transmissíveis/métodos , Doenças Transmissíveis/epidemiologia , Farmacorresistência Bacteriana , Promoção da Saúde/organização & administração , Vigilância de Evento Sentinela , Antibacterianos/uso terapêutico , Doenças Transmissíveis/tratamento farmacológico , Humanos , Programas Nacionais de Saúde/organização & administração , África do SulAssuntos
Doenças Transmissíveis/epidemiologia , Farmacorresistência Bacteriana , Controle de Infecções/métodos , Serviços Preventivos de Saúde/organização & administração , Antibacterianos/uso terapêutico , Doenças Transmissíveis/tratamento farmacológico , Humanos , Programas Nacionais de Saúde/organização & administração , África do SulRESUMO
Oncolytic adenoviruses are being investigated as potential anti-cancer agents. Selective lytic replication in cancer cells is essential for an effective and safe treatment. In this study, we compared 11 oncolytic adenoviruses in relevant cell cultures to assess their use for treating oral cancer and pre-cancerous lesions. We determined the cytotoxicity of oncolytic adenovirus infection and calculated selectivity indices for cytotoxicity to cancer cells compared with normal oral keratinocytes and fibroblasts. Keratinocytes were very sensitive to wild-type adenovirus serotype 5 (Ad5); 1- to 3-log more than head and neck squamous cell carcinoma (HNSCC) cells. The potencies of oncolytic adenoviruses to kill HNSCC cells within 7 days after infection ranged from approximately 10 times less potent to approximately 10 times more potent than Ad5. The selectivity indices determined on fibroblasts and keratinocytes differed markedly. Two oncolytic adenoviruses were more selective than Ad5 for HNSCC cells compared with fibroblasts; and five viruses showed selective replication on HNSCC cells compared with keratinocytes. Overall, CRAd-S.RGD with E1A driven by the survivin promoter and an infectivity-enhancing capsid modification showed the most favourable cytotoxicity pattern; being very potent in killing HNSCC cells, only slightly less effective than Ad5 in killing pre-neoplastic keratinocytes and the least toxic to normal keratinocytes.
Assuntos
Adenoviridae/genética , Adenoviridae/metabolismo , Vetores Genéticos , Neoplasias Bucais/terapia , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Lesões Pré-Cancerosas/terapia , Carcinoma de Células Escamosas/terapia , Linhagem Celular Tumoral , Sobrevivência Celular , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Humanos , Proteínas Inibidoras de Apoptose/genética , Neoplasias Bucais/patologia , Vírus Oncolíticos/metabolismo , Regiões Promotoras Genéticas/genética , SurvivinaRESUMO
INTRODUCTION: Patients with end-stage renal disease suffer from increased genomic damage and cancer incidence. One possible reason is the accumulation of uremic toxins such as homocysteine (Hcy). Elevated Hcy levels--usually indicative of cardiovascular events--correlated with the genomic damage in cross-sectional studies. Therefore we investigated the genotoxic effects of Hcy in vitro. METHODS: To analyse the genomic damage, micronucleus tests and the comet-assay were performed in L5178Y and HL60 cells. Additionally, the influence of Hcy on cell cycle progression, DNA-cytosine-methylation, oxidative stress levels and on the cellular glutathione content were determined. RESULTS: Low millimolar concentrations of Hcy-induced micronuclei in both cell lines but did not enhance the DNA damage observed with the comet-assay. Cell cycle progression was inhibited in S-phase, while DNA-cytosine-methylation remained unchanged. Furthermore, Hcy protected cells challenged with H(2)O(2) from oxidative stress. This was accompanied by an increased cellular glutathione level. CONCLUSION: Since the genotoxic effect was limited to high Hcy concentrations, a contribution of Hcy to the enhanced genomic damage in end-stage renal disease patients would only be conceivable upon local Hcy accumulation. Whether the detected antioxidant capacity of Hcy is relevant for any situation in patients remains to be elucidated.
Assuntos
Antioxidantes/toxicidade , Homocisteína/toxicidade , Mutagênicos/toxicidade , Animais , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citosina/metabolismo , Metilação de DNA , Camundongos , Estresse OxidativoRESUMO
Cancer of the uterine cervix is the second most common cancer in women worldwide. Currently, cervical screening is based on cytology alone. Because infection with high-risk human papillomavirus types (hrHPVs) is a necessary cause of cervical cancer, it has been postulated that screening might become more efficient when it is based on combined cytology and hrHPV testing. In this review we will discuss the advantages of added HPV tests in cervical cancer screening, as a quality control for false-negative smears, in triage of women with equivocal smears, in follow-up of women treated for CIN3 or cervical cancer and for the detection of cervical adenocarcinoma.
Assuntos
Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/diagnóstico , Esfregaço Vaginal , Adenocarcinoma/diagnóstico , Citodiagnóstico/métodos , Reações Falso-Negativas , Feminino , Humanos , Programas de Rastreamento , Neoplasias do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/prevenção & controle , Displasia do Colo do Útero/diagnóstico , Displasia do Colo do Útero/terapiaRESUMO
OBJECTIVE: In the present study the relationship was evaluated between perioperative inflammation and the postoperative acute phase response in patients undergoing elective coronary artery bypass grafting (CABG) assisted by cardiopulmonary bypass (CPB). CPB circuits contained either non-coated- (UMS), Carmeda- (BPS) or Trillium-coated oxygenators (BAS). METHODS: Prospectively, 71 CABG patients were randomly allocated to one of the oxygenator groups (UMS: n=25, BPS: n=25 and BAS: n=21). Terminal complement complexes (TCC) and elastase were determined in plasma samples collected before, during and after bypass. Secretory phospholipase A2 (sPLA2) and C-reactive protein (CRP) were determined before and after bypass. RESULTS: Demographic, CPB and clinical outcome data were similar for the three groups. TCC and elastase increased during CPB, and decreased thereafter. Significant differences between the groups were present in the levels of TCC at the end of CPB (P=0.002) and at the first (P=0.012) and second (P<0.001) postoperative days, the BPS and BAS groups having reduced levels of TCC compared to the UMS group. Also elastase concentrations differed significantly between the groups at the end of CPB (P<0.001). The postoperative sPLA2 and CRP levels increased in all three groups on the first and second postoperative days, but no significant differences were present between the groups. CONCLUSIONS: Material-induced reduction of the inflammatory response during CPB does not affect the postoperative acute phase response. Thus, in CABG patients this response seems relatively unaffected by the composition and/or biocompatibility of the modern CPB circuit and rather to be evoked by surgical trauma, anesthetics and organ perfusion.
Assuntos
Reação de Fase Aguda/etiologia , Ponte Cardiopulmonar/instrumentação , Ativação do Complemento , Reação de Fase Aguda/imunologia , Adulto , Idoso , Proteína C-Reativa/metabolismo , Materiais Revestidos Biocompatíveis , Ponte de Artéria Coronária , Feminino , Humanos , Período Intraoperatório , Masculino , Pessoa de Meia-Idade , Elastase Pancreática/sangue , Fosfolipases A/sangue , Fosfolipases A2 , Período Pós-Operatório , Estudos Prospectivos , Propriedades de Superfície , TrilliumRESUMO
BACKGROUND: A lateral-flow (LF) device using the new reporter up-converting phosphor technology (UPT) was applied to DNA (hybridization) assays for the detection of specific nucleic acid sequences, thereby aiming to perform the test outside well-equipped laboratories. The methodology reported here is sensitive and provides a rapid alternative for more elaborate gel electrophoresis and Southern blotting. In a preliminary study, it was applied to screen for the presence of human papillomavirus type 16 (HPV16) in a defined series of cervical carcinomas. METHODS: A LF assay was used to capture haptenized DNA molecules and hybrids, which were immunolabeled (before LF) with 400-nm UPT particles. These particles emit visible light after excitation with infrared in a process called up-conversion. Because up-conversion occurs in only the phosphor lattice, autofluorescence of other assay components is virtually nonexistent. RESULTS: The use of the UPT reporter in LF-DNA tests, as compared with colloidal gold, improved the detection limit at least 100-fold. UPT LF-DNA tests were successfully applied to detect (in a blind test) the presence of HPV16 in DNA extracts obtained from cervical carcinomas. Test results matched 100% with previous characterization of these carcinomas. CONCLUSIONS: The use of UPT in LF assays to detect specific nucleic acids provides low attamole-range sensitivity. Hybridization and consecutive detection of PCR-amplified HPV16 sequences were successful in a background of 10 microg of fish-sperm DNA. The sensitivity of UPT detection in these complex mixtures indicates that detection of viral infections without PCR or other amplification technique is achievable.