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BACKGROUND: Computed tomography (CT)-guided percutaneous transthoracic needle biopsy (PTNB) is highly affected by respiratory motion; however, respiratory motion of target nodule during the PTNB and its effect on CT-guided lung biopsy have not been studied. PURPOSE: To investigate the effect of the respiratory motion of pulmonary nodules on CT-guided PTNB. MATERIAL AND METHODS: We retrospectively reviewed the procedural CT scans of 426 pulmonary nodules that underwent PTNB during quiet breathing. Maximal and average respiratory motions were measured using the difference of table position of the targeted nodule between multiple procedural scans. Diagnostic performance, complications, and technical factors of PTNB in nodules with large motion (maximal motion >1â cm) were compared with those in nodules with small motion (≤1â cm). RESULTS: The mean maximal and average respiratory motions between tidal volume breathing were 5.4 ± 4.4 and 2.7 ± 2.6â mm, respectively. Sensitivity and accuracy were 93.1% and 96.1% in nodules with large motion, compared with 94.7% and 95.9% in nodules with small motion, respectively. Respiratory targeting (P < 0.001), needle modulation (P < 0.001), motion artifact of target (P < 0.001), target disappearance from scans (P < 0.001), and number of performed CT scans (P < 0.001) were significantly higher in the large motion group, with no significant difference in radiation dose and complications between the groups. CONCLUSION: The respiratory motion of pulmonary nodules during CT-guided PTNB may cause technical difficulties but does not affect diagnostic performance nor complications associated with PTNB.
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Neoplasias Pulmonares , Nódulos Pulmonares Múltiples , Humanos , Neoplasias Pulmonares/patología , Estudios Retrospectivos , Pulmón/diagnóstico por imagen , Pulmón/patología , Nódulos Pulmonares Múltiples/diagnóstico por imagen , Tomografía Computarizada por Rayos X/métodos , Biopsia con Aguja/métodos , Biopsia Guiada por Imagen/métodos , Radiografía Intervencional/métodosRESUMEN
We present a rare case of diffuse large B-cell lymphoma developed in subcutaneous fat layer of the breast with cardiac involvement. Radiologists should perform an image-guided biopsy for pathologic confirmation of breast lymphomas and avoidance of unnecessary invasive treatment.
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Neoplasias de la Mama , Linfoma de Células B Grandes Difuso , Humanos , Femenino , Linfoma de Células B Grandes Difuso/diagnóstico por imagen , Linfoma de Células B Grandes Difuso/patología , Neoplasias de la Mama/diagnóstico por imagen , Neoplasias de la Mama/patología , Grasa Subcutánea/diagnóstico por imagen , Grasa Subcutánea/patologíaRESUMEN
We present a rare case of male axillary accessory breast cancer, which is extremely rare and is indistinguishable from lymphadenopathy and other malignancies, such as lymphoma and skin-derived tumors. Clinicians should consider accessory breast cancer in the differential diagnosis even in men, particularly in those who present with superficially located tumors with adjacent accessory breast tissue.
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Enfermedades de la Mama , Neoplasias de la Mama Masculina , Neoplasias de la Mama , Coristoma , Axila , Mama/diagnóstico por imagen , Mama/patología , Enfermedades de la Mama/patología , Neoplasias de la Mama/patología , Neoplasias de la Mama Masculina/diagnóstico por imagen , Neoplasias de la Mama Masculina/patología , Coristoma/diagnóstico por imagen , Humanos , MasculinoRESUMEN
PURPOSE: To evaluate the diagnostic performance, image quality, artifacts, and radiation doses of virtual non-contrast (VNC) images, relative to true non-contrast (TNC) images, in patients with surgically clipped aneurysms. METHODS: Seventy-six patients who underwent unenhanced computed tomography (CT) and dual-energy (DE)-CT angiography after surgical clipping of 85 intracranial aneurysms were included in the study. Diagnostic performances of VNC and TNC images were compared with respect to sensitivity, specificity, and positive and negative predictive values. The results of quantitative and qualitative analyses were compared between VNC and TNC images. Radiation doses were also compared between VNC and TNC images. RESULTS: Diagnostic performance of VNC images was lower than that of TNC images; however, addition of contrast-enhanced images improved diagnostic performance. Image noise and mean attenuation of VNC images were significantly higher than those of TNC images in the centrum semiovale, cerebellum, and pons. The quality of VNC images was excellent or sufficient (85.5% for observer 1, 85.5% for observer 2), and complete acceptance of VNC images was achieved in 64.5% for observer 1 and in 71.0% for observer 2; however, the addition of contrast-enhanced images increased the level of acceptance (92.0% for observer 1, 90.9% for observer 2). Clip artifacts were significantly lower in VNC images than in TNC images. CT dose index, dose-length product, and effective dose were significantly lower without TNC images. CONCLUSION: VNC images showed lower diagnostic performance and image quality, and higher image noise than TNC images; however, VNC images could reduce clip artifacts and radiation doses.
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Angiografía por Tomografía Computarizada/métodos , Aneurisma Intracraneal/diagnóstico por imagen , Adulto , Anciano , Anciano de 80 o más Años , Artefactos , Medios de Contraste , Femenino , Humanos , Aneurisma Intracraneal/cirugía , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Sensibilidad y EspecificidadRESUMEN
BACKGROUND This study aimed to assess the utility and characteristics of preoperative ultrasonography (US) in patients transferred to referral hospitals from local clinics with a diagnosis of malignancy on US-guided fine-needle aspiration cytology of thyroid nodules. MATERIAL AND METHODS From January 2018 to June 2018, 109 transferred patients underwent preoperative US in our hospital for suspected thyroid malignancy on cytological analysis after US-guided fine-needle aspiration of thyroid nodules in local clinics. Preoperative US was performed by a single radiologist in all patients. Among them, 6 were excluded from the study because of refusal of thyroid surgery. Preoperative US and histopathological results were compared in all patients. RESULTS After thyroid surgery, pathological examination revealed papillary thyroid carcinoma (PTC) (n=98), follicular adenoma (n=1), and nodular hyperplasia (n=4). Of the 103 patients, 91 exhibited suspicious US findings on the preoperative US, whereas 12 did not. In the 91 patients with suspicious US findings, PTC (n=90) and follicular adenoma (n=1) were confirmed after thyroid surgery. In the 12 patients with no suspicious US findings, PTC (n=8) and nodular hyperplasia (n=4) were confirmed after thyroid surgery. On repeat analysis of the cytological slides of the 4 nodular hyperplasia cases from the local clinics, Bethesda category II (n=1) and III (n=3) were determined. CONCLUSIONS In the transferred patients with a malignant cytology, preoperative US might be helpful to detect false-positive cytology cases.
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Cuidados Preoperatorios , Utilización de Procedimientos y Técnicas , Glándula Tiroides/diagnóstico por imagen , Glándula Tiroides/patología , Nódulo Tiroideo/diagnóstico por imagen , Nódulo Tiroideo/patología , Ultrasonografía , Adolescente , Adulto , Anciano , Biopsia con Aguja Fina , Carcinoma Papilar/diagnóstico por imagen , Carcinoma Papilar/patología , Carcinoma Papilar/cirugía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Transferencia de Pacientes , Estudios Retrospectivos , Glándula Tiroides/cirugía , Neoplasias de la Tiroides/diagnóstico por imagen , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/cirugía , Nódulo Tiroideo/cirugía , Adulto JovenRESUMEN
BACKGROUND This study aimed to evaluate the prevalence of thyroglossal duct cysts (TGDCs) on ultrasonography (US) and US features of TGDCs in adults, and to assess whether the prevalence or size of TGDCs increases after radioactive iodine ablation (RIA). MATERIAL AND METHODS Between July and December 2018, 2820 patients underwent thyroid or neck US examination, performed by 2 radiologists, at our center. On the basis of real-time US, the presence or absence of TGDCs was prospectively investigated by 2 radiologists. Among the 2820 patients, 54 patients who were <19 years of age or had a radiation therapy history to the neck were excluded. Eventually, 2766 patients were included. RESULTS Of the 2766 patients, 160 (5.8%) showed a TGDC on US. The mean size of TGDCs in RIA history (+) (n=36) and RIA history (-) (n=124) groups was 0.92±0.41 cm and 0.86±0.45 cm, respectively. There was no significant difference in size of TGDCs between RIA history (+) and RIA history (-) groups (p=0.684). Between the TGDC (+) and TGDC (-) groups, there was no significant difference in patient age, gender, reason for thyroid/neck US, type of thyroid surgery, and session number and application/no application of RIA (p>0.05). The prevalence rate of TGDCs in radiologist A and B was 4.9% (70/1427) and 6.7% (90/1339), respectively. TGDCs were more common in the suprahyoid neck, and the common shapes of TGDCs were flat-to-ovoid and round. CONCLUSIONS RIA may not be associated with the prevalence or enlargement of TGDCs.
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Quiste Tirogloso/diagnóstico por imagen , Quiste Tirogloso/radioterapia , Técnicas de Ablación/métodos , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma Papilar , Niño , Preescolar , Femenino , Humanos , Lactante , Yodo , Radioisótopos de Yodo , Masculino , Persona de Mediana Edad , Prevalencia , Estudios Retrospectivos , Glándula Tiroides , Neoplasias de la Tiroides , Tomografía Computarizada por Rayos X , Ultrasonografía/métodosRESUMEN
BACKGROUND: Background parenchymal enhancement (BPE) on dynamic contrast-enhanced (DCE)-MRI has been associated with breast cancer risk, both based on qualitative and quantitative assessments. PURPOSE: To investigate whether BPE of the contralateral breast on preoperative DCE-MRI is associated with therapy outcome in ER-positive, HER2-negative, node-negative invasive breast cancer. STUDY TYPE: Retrospective. POPULATION: In all, 289 patients with unilateral ER-positive, HER2-negative, node-negative breast cancer larger than 5 mm. FIELD STRENGTH/SEQUENCE: 3T, T1 -weighted DCE sequence. ASSESSMENT: BPE of the contralateral breast was assessed qualitatively by two dedicated radiologists and quantitatively (using region-of-interest and automatic breast segmentation). STATISTICAL TESTS: Cox regression analysis was used to determine associations with recurrence-free survival (RFS) and distant metastasis-free survival (DFS). Interobserver variability for parenchymal enhancement was assessed using kappa statistics and intraclass correlation coefficient (ICC). RESULTS: The median follow-up time was 75.8 months. Multivariate analysis showed receipt of total mastectomy (hazard ratio [HR]: 5.497) and high Ki-67 expression level (HR: 5.956) were independent factors associated with worse RFS (P < 0.05). Only a high Ki-67 expression level was associated with worse DFS (HR: 3.571, P = 0.045). BPE assessments were not associated with outcome (RFS [qualitative BPE: P = 0.75, 0.92 for readers 1 and 2; quantitative BPE: P = 0.38-0.99], DFS, [qualitative BPE: P = 0.41, 0.16 for readers 1 and 2; quantitative BPE: P = 0.68-0.99]). For interobserver variability, there was good agreement between qualitative (κ = 0.700) and good to perfect agreement for most quantitative parameters of BPE. DATA CONCLUSION: Contralateral BPE showed no association with survival outcome in patients with ER-positive, HER2-negative, node-negative invasive breast cancer. A high Ki-67 expression level was associated with both worse recurrence-free and distant metastasis-free survival. LEVEL OF EVIDENCE: 3 Technical Efficacy: Stage 4 J. Magn. Reson. Imaging 2018;48:1678-1689.
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Neoplasias de la Mama/diagnóstico por imagen , Neoplasias de la Mama/metabolismo , Medios de Contraste/química , Imagen por Resonancia Magnética , Adulto , Anciano , Anciano de 80 o más Años , Mama/diagnóstico por imagen , Mama/patología , Supervivencia sin Enfermedad , Receptor alfa de Estrógeno/metabolismo , Femenino , Estudios de Seguimiento , Humanos , Metástasis Linfática , Mastectomía , Ciclo Menstrual , Persona de Mediana Edad , Análisis Multivariante , Invasividad Neoplásica , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/patología , Variaciones Dependientes del Observador , Modelos de Riesgos Proporcionales , Receptores de Progesterona/metabolismo , Análisis de Regresión , Estudios RetrospectivosRESUMEN
OBJECTIVE: It is uncertain whether papillary thyroid carcinomas (PTCs) of the same subtype display similar sonographic features. This retrospective analysis of pre-operative sonographic and pathologic findings aimed to assess whether PTCs of the same subtype share sonographic features. METHODS: Before undergoing thyroid surgery, 137 patients underwent ultrasound (US) examination. A single radiologist used a picture archiving and communication system and pathologic reports to investigate all sonographic features of the largest and second largest PTCs. Additionally, the radiologist evaluated the similarity of sonographic features between primary (largest), secondary (second largest), and daughter (secondary with same subtype as the primary) PTCs. RESULTS: Of the 137 PTC patients, 48 (35.0%) had multiple PTCs; however, 5 had no US images of the secondary PTC. Of the 43 secondary PTCs with US images, 9 (20.9%) secondary PTCs were of a different subtype than the primary PTC and revealed sonographic features that differed from those of the primary PTC. Of the 48 patients with multiple PTCs, the subtype was the same in the primary and secondary PTCs in 34 (70.8%) patients. Of the 34 daughter PTCs, 32 (94.1%) had sonographic features similar to those of the primary PTC, whereas 2 (5.9%) showed different sonographic features than the primary PTC. There was no significant difference between primary and daughter PTCs in the size, location, sonographic features, or Korean Thyroid Imaging Reporting and Data System category ( P>.05). CONCLUSION: Daughter PTCs show similar sonographic features as the primary PTC. ABBREVIATIONS: K-TIRADS = Korean Thyroid Imaging Reporting and Data System; PTC = papillary thyroid carcinoma; US = ultrasound.
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Carcinoma Papilar/diagnóstico por imagen , Neoplasias de la Tiroides/diagnóstico por imagen , Adolescente , Adulto , Anciano , Carcinoma Papilar/patología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Cáncer Papilar Tiroideo , Neoplasias de la Tiroides/patología , Ultrasonografía , Adulto JovenRESUMEN
In this genomic era, the ability to assay multiple genomic hot spots that have strong clinical implications is greatly desired. Conventional PCR-based methods suffer from frequent false-positive detections, particularly when a multiplex analysis is desirable. As an alternative to the error-prone conventional methods, multiplex ligase-based genotyping methods combined with CE have a strong potential. In this review, both previously developed methods and emerging methods are described to reveal the specificity, sensitivity, and simplicity of the ligase-based methods. For each step (ligation, amplification, and separation), the principles of several alternative methods are discussed along with their applications to explore the future development of ligase-based diagnostic methods.
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Electroforesis Capilar/métodos , Técnicas de Genotipaje/métodos , Reacción en Cadena de la Polimerasa/métodos , Animales , Humanos , Sensibilidad y EspecificidadRESUMEN
A simple and precise diagnostic method for spinal muscular atrophy (SMA) using high-resolution CE-based single-strand conformation polymorphism (CE-SSCP) was developed in this study. SMA is a common genetic disorder caused by an abnormality in the relative copy numbers of SMN1 and its centromeric copy SMN2, which differ only in two nucleotides, namely at exons 7 and 8. Therefore, the precise discrimination of the differences in sequence as well as their relative quantities is crucial for the diagnosis of SMA. Multiplex ligation-dependent probe amplification and sequence-sensitive DNA separation using hydroxyethyl cellulose and hydroxypropyl cellulose blended polymer matrix are currently the available methods used in the diagnosis of SMA. However, these methods are limited by their extended hybridization step and low resolution. In this study, the simultaneous discrimination of SMN exons 7 and 8 was successfully demonstrated using high-resolution CE-SSCP. Unlike the previously reported alternative method, single base differing amplicons were baseline-separated because of its extraordinary resolution, thus providing accurate and precise quantification of each paralog.
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Electroforesis Capilar/métodos , Dosificación de Gen , Atrofia Muscular Espinal/genética , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Humanos , Masculino , Polimorfismo de Nucleótido Simple , Polimorfismo Conformacional Retorcido-SimpleRESUMEN
Accuracy, simplicity, and cost-effectiveness are the most important criteria for a genotyping method for SNPs compatible with clinical use. One method developed for SNP genotyping, ligase-based discrimination, is considered the simplest for clinical diagnosis. However, multiplex assays using this method are limited by the detection method. Although CE has been introduced as an alternative to error prone microarray-based detection, the design process and multiplex assay procedure are complicated because of the DNA size-dependent separation principle. In this study, we developed a simple and accurate multiplex genotyping method using reaction condition-optimized ligation and high-resolution CE-based SSCP. With this high-resolution CE-SSCP system, we are able to use similar-sized probes, thereby eliminating the complex probe design step and simplifying the optimization process. We found that this method could accurately discriminate single-base mismatches in SNPs of the tp53 gene, used as targets for multiplex detection.
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Disparidad de Par Base , Polimorfismo de Nucleótido Simple , Polimorfismo Conformacional Retorcido-Simple , Secuencia de Bases , ADN Ligasas/metabolismo , Electroforesis Capilar/métodos , Genotipo , Humanos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Proteína p53 Supresora de Tumor/genéticaRESUMEN
CE-SSCP analysis is a well-established DNA separation method that is based on variations in mobility caused by sequence-induced differences in the conformation of single-stranded DNA. The resolution of CE-SSCP analysis was improved by using a Pluronic polymer matrix, and it has been successfully applied in various genetic analyses. Because the Pluronic polymer forms a micellar cubic structure in the capillary, it provides a stable internal structure for high-resolution CE-SSCP analysis. We hypothesized that formation of micellar cubic structure is influenced by the end hydroxyl group of the Pluronic polymer, which affords structural stability through hydrogen bonding. To test this hypothesis, the hydroxyl group was halogenated to eliminate the hydrogen bonding without disturbing the polarity of polymer matrix. CE-SSCP resolution of two DNA fragments with a single base difference was significantly worse in the halogenated polymer matrices due to band broadening. The viscoelastic properties of control (which has hydroxyl group), chlorinated, and brominated F108 solution upon heating were also investigated by rheological experiments, and we found that gelation was significantly associated with resolution. In this series of experiments, the effect of the hydroxyl group in Pluronic polymer matrix on separation resolution of CE-SSCP analysis was demonstrated.
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ADN/aislamiento & purificación , Poloxámero/química , ADN/análisis , ADN/química , Electroforesis Capilar/métodos , Halogenación , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , TemperaturaRESUMEN
Antibody-conjugated magnetic nanoparticles (Ab-MNPs) have potential in pathogen detection because they allow target cells to be easily separated from complex sample matrices. However, the sensitivity and specificity of pathogen capture by Ab-MNPs generally vary according to the types of MNPs, antibodies, and sample matrices, as well as preparation methods, including immobilization. Therefore, achieving a reproducible analysis utilizing Ab-MNPs as a pathogen detection method requires accurate characterization of Ab-MNP capture ability and standardization of all handling processes. In this study, we used high-resolution CE-single strand conformational polymorphism coupled with a stuffer-free multiplex ligation-dependent probe amplification system to characterize Ab-MNPs. The capture ability of Ab-MNPs targeting Salmonella enteritidis and nine pathogens, including S. enteritidis, was analyzed in phosphate buffer and milk. The effect of storage conditions on the stability of Ab-MNPs was also assessed. The results showed that the stuffer-free multiplex ligation-dependent probe amplification system has the potential to serve as a standard characterization method for Ab-MNPs. Moreover, the precise characterization of Ab-MNPs facilitated robust pathogen detection in various applications.
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Anticuerpos/química , Bacterias/genética , Técnicas de Tipificación Bacteriana/métodos , Nanopartículas de Magnetita/química , Reacción en Cadena de la Polimerasa Multiplex/métodos , Nanoconjugados/química , Animales , Bacterias/clasificación , Bacterias/aislamiento & purificación , Electroforesis Capilar , Límite de Detección , Carne/microbiología , Leche/microbiología , Polimorfismo Conformacional Retorcido-SimpleRESUMEN
Lower respiratory tract infection is one of the most common infectious diseases. However, conventional methods for detecting infectious pathogens are time-consuming, and generally have a limited impact on early therapeutic decisions. We previously reported a rapid and sensitive method for detecting such pathogens using stuffer-free multiplex ligation-dependent probe amplification coupled with high-resolution CE-SSCP. In this study, we report an application of this method to the detection of respiratory pathogens. As originally configured, this method was capable of simultaneously detecting seven bacterial species responsible for lower respiratory tract infections, but its detection limit and assay time were insufficient to provide useful information for early therapeutic decisions. To improve sensitivity and shorten assay time, we added a target-specific preamplification step, improving the detection limit from 50 pg of genomic DNA to 500 fg. We further decreased time requirements by optimizing the hybridization step, enabling the entire assay to be completed within 7 h while maintaining the same detection limit. Taken together, these improvements enable the rapid detection of infectious doses of pathogens (i.e. a few dozen cells), establishing the strong potential of the refined method, particularly for aiding early treatment decisions.
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Bacterias/genética , Tipificación Molecular/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , Infecciones del Sistema Respiratorio/microbiología , Bacterias/clasificación , Infecciones Bacterianas/microbiología , Sondas de ADN , ADN Bacteriano/análisis , ADN Bacteriano/genética , Electroforesis Capilar/métodos , Humanos , Límite de Detección , Polimorfismo Conformacional Retorcido-Simple , Esputo/microbiologíaRESUMEN
Breast lymphomas are rare, malignant breast neoplasms with a heterogeneous pattern of clinical symptoms. Burkitt's lymphoma is a rare, highly aggressive, and rapidly growing B-cell non-Hodgkin lymphoma. We report about a 27-year-old woman diagnosed as having secondary breast Burkitt's lymphoma, probably originating from the stomach, with multiple distant metastases. Breast ultrasonography revealed multiple, variable sized, heterogeneous masses with posterior acoustic enhancement and echogenic rims. These imaging findings may sometimes overlap with those of other breast malignancies. However, unlike other breast malignancies, lymphoma can be diagnosed by biopsy and does not require surgical excision. To avoid unnecessary treatment, radiologists and clinicians should be aware of the characteristic imaging features of breast lymphomas.
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Tuberculous pericarditis is an extrapulmonary manifestation of tuberculosis that is most commonly associated with pericardial thickening, effusion, and calcification. We present a case of tuberculous pericarditis mimicking a malignant pericardial tumor in a 77-year-old male. CT revealed an irregular and nodular pericardial thickening. MRI revealed high signal intensity on T1-weighted fat-suppressed images and peripheral rim enhancement after gadolinium administration. MRI can be helpful in determining the differential diagnoses in cases of tuberculous pericarditis with nonspecific imaging findings.
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This study investigated the effect of temperature on the aspect-ratio etching of SiO2 in CF4/H2/Ar plasma using patterned samples of a 200 nm trench in a low-temperature reactive-ion etching system. Lower temperatures resulted in higher etch rates and aspect ratios for SiO2. However, the plasma property was constant with the chuck temperature, indicated by the line intensity ratio from optical emission spectroscopy monitoring of the plasma. The variables obtained from the characterization of the etched profile for the 200 nm trench after etching were analyzed as a function of temperature. A reduction in the necking ratio affected the etch rate and aspect ratio of SiO2. The etching mechanism of the aspect ratio etching of SiO2 was discussed based on the results of the surface composition at necking via energy-dispersive X-ray spectroscopy with temperature. The results suggested that the neutral species reaching the etch front of SiO2 had a low sticking coefficient. The bowing ratio decreased with lowering temperature, indicating the presence of directional ions during etching. Therefore, a lower temperature for the aspect ratio etching of SiO2 could achieve a faster etch rate and a higher aspect ratio of SiO2 via the reduction of necking than higher temperatures.
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In systems biological studies, precise expression profiling of functionally important gene sets is crucial. Real-time polymerase chain reaction is generally used for this purpose. Despite its widespread acceptance, however, this method is not suitable for multiplex analysis, resulting in an inefficient assay process. One alternative technology in the spotlight is multiplex ligation-dependent probe amplification (MLPA). But MLPA depends on length-based discrimination of amplified products, which complicates probe design and compromises analysis results. Here, we devised a variation of MLPA that utilizes conformation-sensitive capillary electrophoresis, and demonstrated the simplicity of the probe-design process and improved precision of the assay in analyses of 33 Escherichia coli metabolic genes and 16 Caenorhabditis elegans longevity-related genes. The results showed that relative expression could be quantitatively measured over a relevant dynamic range by using similar-sized probes. Importantly, the improved precision compared to conventional MLPA promises a wider application of this method for various biological systems.
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Caenorhabditis elegans/genética , Escherichia coli/genética , Reacción en Cadena de la Polimerasa Multiplex , Animales , Caenorhabditis elegans/metabolismo , Electroforesis Capilar , Escherichia coli/metabolismoRESUMEN
The success of antimicrobial therapy depends on effective prescription of antibiotics. Assessment of clinical isolates using rapid antimicrobial susceptibility tests allows effective microbiological therapy to be commenced in a timely manner. However, conventional antimicrobial susceptibility testing is time-consuming and laborious. In the present study, we employed stuffer-free multiplex ligation-dependent probe amplification (MLPA) coupled with analysis of single-strand conformation polymorphisms, via high-resolution CE, to develop a multiplex antibiotic susceptibility test. Using this method, parallel analysis of specific genetic markers was employed to determine minimal inhibitory concentration values. The values derived using the stuffer-free MLPA method agreed with those estimated using a conventional broth dilution method. These findings indicate that the stuffer-free MLPA-based approach is a viable alternative to the conventional method.
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Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Electroforesis Capilar/métodos , Pruebas de Sensibilidad Microbiana/métodos , Reacción en Cadena de la Polimerasa Multiplex/métodos , Bacterias/genética , Bacterias/aislamiento & purificación , ADN Bacteriano/análisis , ADN Bacteriano/genética , Polimorfismo Conformacional Retorcido-SimpleRESUMEN
Pluronic F108 block copolymers have shown a great promise to achieve the desirable high resolution in the conformation-sensitive separation of ssDNA using CE-SSCP. However, fundamental understanding of the structures and properties of Pluronic matrix affecting the resolution is still limited. Unlike conventional gel-forming homopolymers, Pluronic F108 block copolymers are amphiphilic macromolecules consisting of poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) triblock copolymers, which are capable of forming a highly ordered micellar structure in aqueous solution. In this study, we have performed a series of experiments by blending different types of Pluronic polymers to control the formation of micelles and to study the correlation between separation and rheological characteristics of Pluronic gels affecting the resolution of CE-SSCP. Our experiments have been specifically designed to elucidate how the micellar structure affects the resolution of CE-SSCP upon altering the size uniformity and constituent homogeneity of the micelles. Our results suggest that uniformly sized micelle packing is the primary structural feature of Pluronic gel matrix for the high-resolution separation, while the size and constituent of the micelle themselves need to be considered as secondary factors.