Applications of microchip electrophoresis and capillary electrophoresis for screening FLT3-ITD gene mutation in acute myeloid leukemia / 中国实验血液学杂志

The purpose of the present study was to compare the reliability of microchip electrophoresis and capillary electrophoresis for screening FLT3-ITD gene mutation in acute myeloid leukemia. The FLT3-ITD mutation in the genomic DNA samples from 214 untreated AML patients were separately detected by PCR-microchip electrophoresis and PCR-capillary electrophoresis, then the DNA direct sequencing analysis was carried out. The results from PCR-microchip electrophoresis showed that there were 151 FLT3-ITD mutation negative, 58 FLT3-ITD mutation positive (58/214, 27.1%) and 5 FLT3-ITD mutation doubtful positive (5/214, 2.3%), while the outcomes from PCR-capillary electrophoresis displayed that there were 147 FLT3-ITD mutation negative and 67 FLT3-ITD mutation positive (67/214, 31.3%) without doubtful positive. In the 67 FLT3-ITD mutation positive samples detected by using PCR-capillary electrophoresis, 4 samples were detected as the negative while 5 samples were measured as the doubtful positive by using PCR-microchip electrophoresis. The followed sequencing analysis demonstrated that the above 9 samples were all FLT3-ITD mutation positive, indicating that PCR-capillary electrophoresis was more accurate and sensitive in screening the FLT3-ITD mutation, although statistic analysis showed that there were no significant differences in the detected results between PCR-microchip electrophoresis and PCR-capillary electrophoresis groups (Pearson Chi-squared Test, P > 0.05). It is concluded that both PCR-microchip electrophoresis and PCR-capillary electrophoresis were convenient and fast for screening FLT3-ITD mutation, but the accuracy of PCR-microchip electrophoresis awaits further improvement.

Expression and clinical significance of secretory leucocyte protease inhibitor in colon carcinoma / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the expression of secretory leucocyte protease inhibitor (SLPI) in colon cancer and their clinical significance.</p><p><b>METHODS</b>Immunohistochemistry was performed to detect the SLPI expression in colon cancer tissue microarray. The expression of SLPI was scored by two pathologists and was analyzed using Χ(2) test to explore its influence on the pathologic characteristics of colon carcinoma.</p><p><b>RESULTS</b>SLPI was up-regulated in colon cancer tissue compared to normal mucosa. Overexpression of SLPI protein was correlated with differentiation grade (low differentiation: 42.1% vs 57.9%; moderate/well differentiation: 2.3% vs 97.7%, TNM stages(III-IV:29.4% vs 70.6%;I-II:3.1% vs 96.9%), lymph node metastasis (28.6% vs 71.4%) and distant metastasis (84.6% vs 15.4%), but not with patient age or sex.</p><p><b>CONCLUSION</b>SLPI overexpression correlates with aggressive pathologic characteristics of colon cancer and it may server as prognostic factor of colon cancer patients. Further research will be carried out to verify whether SLPI can become a new target for colon cancer treatment.</p>

Comparison of the DNA Preservation in Neutral-Buffered Formalin Fixed Paraffin-Embedded Tissue and in Non-Buffered Formalin Fixed Paraffin-Embedded Tissue

BACKGROUND: The preservation of optimized DNA and its extraction from formalin-fixed, paraffin-embedded (FFPE) tissues are important issues. There has been some doubt over whether 10% neutral-buffered formalin is an ideal fixation solution for DNA preservation over non-buffered formalin, as conventionally recommended. In this study, the correlation between the efficiency of DNA extraction from FFPE tissues and buffered formalin was evaluated. METHODS: Several tissues with same conditions except fixatives were fixed in four different formalin solution groups and were routinely processed as paraffin-embedding protocols. DNAs were extracted from four different FFPE tissues that were stored for over 3 months and over 9 months. The quantity and quality of the DNAs were assessed with a NanoDrop ND-1000 spectrophotometer, and the polymerase chain reaction (PCR) amplification and degradation were analyzed via microchip electrophoresis. KRAS mutation analysis and microsatellite instability (BAT25) PCR were performed with each sample. RESULTS: The results showed no remarkable difference in the four groups. CONCLUSIONS: The study findings demonstrate that DNA preservation is fairly unaffected by a neutral buffer where there is short formalin manufacture period and an adequate formalin fixation time before embedding in paraffin.

Bacterial handling under the influence of non-uniform electric fields: dielectrophoretic and electrohydrodynamic effects

This paper describes the modeling and experimental verification of a castellated microelectrode array intended tohandle biocells, based on common dielectrophoresis. The proposed microsystem was developed employing platinumelectrodes deposited by lift-off, silicon micromachining, and photoresin patterning techniques. Having fabricated the microdevice it was tested employing Escherichia coli as bioparticle model. Positive dielectrophoresis could be verified with the selected cells for frequencies above 100 kHz, and electrohydrodynamic effects were observed as the dominant phenomena when working at lower frequencies. As a result, negative dielectrophoresis could not be observed because its occurrence overlaps with electrohydrodynamic effects; i.e. the viscous drag force acting on the particles is greater than the dielectrophoretic force at frequencies where negative dielectrophoresis should occur. The experiments illustrate the convenience of this kind of microdevices to micro handling biological objects, opening the possibility for using these microarrays with other bioparticles. Additionally, liquid motion as a result of electrohydrodynamic effects must be taken into account when designing bioparticle micromanipulators, and could be used as mechanism to clean the electrode surfaces, that is one of the most important problems related to this kind of devices.

Este artigo descreve a modelagem e teste experimental de uma rede de microeletrodos em cremalheira cujo objetivo é o manuseio de células biológicas, com base em dieletroforese comum. O microsistema proposto foi desenvolvido empregando eletrodos de platina depositados por técnicas de 'lift-off', micro-usinagem em silício e litografia com foto-resina. Uma vez fabricado o microdispositivo, este foi testado utilizando a Escherichia coli como modelo de biopartículas. Dieletroforese positiva pode ser observada com as células selecionadas para freqüências acima de 100kHz, e efeitos eletro-hidrodinâmicos foram observados como o fenômeno dominante para menores freqüências. Como resultado, a dieletroforese negativa não pode ser observada pois sua ocorrência se sobrepõe a efeitos eletro-hidrodinâmicos; i.e. a força de arraste viscoso atuando sobre as partículas é superior à força dieletroforética para freqüências em que a dieletroforese negativa deveria ocorrer. Os experimentos ilustram a conveniência deste tipo de micro-dispositivo para o micromanuseio de objetos biológicos, abrindo a possibilidade de uso destas micro-redes com outras partículas biológicas. Além disto, o movimento líquido como resultado dos efeitos eletro-hidrodinâmicos deve ser levado em conta ao se desenhar micromanipuladores de partículas biológicas, e pode ser utilizado como mecanismo para limpar as superfícies dos eletrodos, que é um dos problemas mais importantes relacionados a este tipo de dispositivo.

The relation between Escherichia coli surface functional groups' composition and their physicochemical properties

Escherichia coli surface characteristics including hydrophobicity, electrophoretic mobility and surface functional groups' composition were investigated. These characteristics were determined respectively by water contact angle measurements, microelectrophoresis and x-ray photoelectron spectroscopy (XPS). The relation between the physicochemical properties and functional groups' composition was also examined. The electrophoretic mobility at pH 7 appeared to be governed on the cell surface by the (O=C) functional groups. The cell surface's hydrophilicity was associated with high levels of (C-(O.N)) and (OH-(C-O-C)) functional groups, whereas the cell surface's hydrophobicity was associated with (C-(C,H)) functional groups.

Características de superfície de Escherichia coli, tais como hidrofobicidade, mobilidade eletroforética e composição dos grupos funcionais de superfície, foram estudadas. Essas características foram determinadas por medidas de angulo de contato com água, microeletroforese e espectroscopia fotoeletrônica de raio-X (XPS), respectivamente. A relação entre as propriedades fisicoquímicas e a composição dos grupos funcionais foi também examinada. Aparentemente, a mobilidade eletroforética em pH 7 é controlada na superfície celular pelos grupos funcionais (O=C). A hidrofilicidade da superfície celular estava associada com altos níveis dos grupos funcionais [C-(O.N)] e [OH-(C-O-C)], enquanto a hidrofobicidade estava associada com os grupos funcionais [C-(C,H)].

Roles of ACh receptors in the effects of corticosterone on presympathetic neurons in the ventrolateral medulla of rats / 中国应用生理学杂志

<p><b>AIM</b>To investigate the roles of acetylcholine (ACh) receptors in the rapid effects of corticosterone (CORT) on the presympathetic neurons in the rostral ventrolateral medulla (RVLM) of rats, and study the non-genomic mechanism of glucocorticoid (GC) in the integration of sympathetic cardiovascular activity.</p><p><b>METHODS</b>The effects of microelectrophoresis of CORT on the discharge of the presympathetic neurons in the RVLM were observed by extracellular recording in urethane-anaesthetized rats. The responses of atropine (a blocker for M type of ACh receptor, ATR), d-tubocurarine (a blocker for N1 type of ACh receptor, d-TC) and hexamethonium (a blocker for N2 type of ACh receptor, C6) to the effects of CORT on the presympathetic neurons were investigated respectively.</p><p><b>RESULTS</b>Totally 33 presympathetic neurons in the RVLM were recorded. Among them the firing rate of 25 (76%) presympathetic neurons was increased by microelectrophoresis of CORT. The effects of CORT were also positively correlated with the currents. In the other 8 presympathetic neurons, had was shown no effect after microelectrophoresis of CORT. In 10 presympathetic neurons, which discharge was increased by CORT, microelectrophoresis of ATR decreased the firing rate of these presympathetic neurons (P < 0.05), and did not fully block the excitatory effect induced by CORT. In both 7 and 6 presympathetic neurons, application of d-TC and C6 had no effect on these neurons respectively, and did not fully block the excitatory effect induced by CORT.</p><p><b>CONCLUSION</b>CORT had rapid excitatory effects on the presympathetic neurons in the RVLM, which effect might be independent on ACh receptors.</p>