AIM: To detect the new serum biomarkers for colorectal cancer (CRC) by serum protein profiling with surfaceenhanced laser desorption ionisation - time of flight mass spectrometry (SELDI-TOF MS). METHODS: Two ind...AIM: To detect the new serum biomarkers for colorectal cancer (CRC) by serum protein profiling with surfaceenhanced laser desorption ionisation - time of flight mass spectrometry (SELDI-TOF MS). METHODS: Two independent serum sample sets were analysed separately with the ProteinChip technology (set A: 40 CRC + 49 healthy controls; set B: 37 CRC + 31 healthy controls), using chips with a weak cation exchange moiety and buffer pH 5. Discriminative power of differentially expressed proteins was assessed with a classification tree algorithm. Sensitivities and specificities of the generated classification trees were obtained by blindly applying data from set A to the generated trees from set B and vice versa. CRC serum protein profiles were also compared with those from breast, ovarian, prostate, and non-small cell lung cancer. RESULTS: Mass-to-charge ratios (m/z) 3.1×10^3, 3.3× 10^3, 4.5×10^3, 6.6×10^3 and 28×10^3 were used as classitiers in the best-performing classification trees. Tree sensitivities and specificities were between 65% and 90%.Host of these discriminative m/z values were also different in the other tumour types investigated. M/z 3.3× 10^3, main classifier in most trees, was a doubly charged form of the 6.6× 10^3-Da protein. The latter was identified as apolipoprotein C-I. M/z 3.1×10^3 was identified as an N-terminal fragment of albumin, and m/z 28× 10^3 as apolipoprotein A-I. CONCLUSION: SELDI-TOF MS followed by classification tree pattern analysis is a suitable technique for finding new serum markers for CRC. Biomarkers can be identified and reproducibly detected in independent sample sets with high sensitivities and specificities. Although not specific for CRC, these biomarkers have a potential role in disease and treatment monitoring.展开更多
Objective: To establish a sensitive and specific system for genotoxicity detection in vivo. Methods: Endogenous p53 tracer vector p53RE was constructed by using green fluorescent protein (GFP) as a reporter to trace p...Objective: To establish a sensitive and specific system for genotoxicity detection in vivo. Methods: Endogenous p53 tracer vector p53RE was constructed by using green fluorescent protein (GFP) as a reporter to trace p53 transcriptional activity under the control of base SV40 early promoter. The tracer cells 3T3-REG were established by transfecting NIH3T3 cells with tracer vector and treated with ultraviolet, H2O2 and adriamycin to induce DNA damage and the subsequent endogenous p53 expression. The GFP expression and its green fluorescence in the tracer cells were observed and measured with fluorescent microscope and FACS. Results: The GFP expression increased rapidly after various treatment and reached the maximum 1 h later, and decreased gradually afterwards. FACS analysis showed that GFP expression in 3T3-REG tracer cells was consistent with the concentration of H2O2, while that in 3T3-SVG cells, which were transfected with control vector pSV-GFP. hardly increased in response to the treatment. Conclusion: GFP tracing p53 transcriptional activity is a sensitive and specific method for genotoxicity detection.展开更多
To compare mid-infrared(MIR)and near-infrared(NIR)spectroscopies for the determination of the fat and protein contents in milk,the same sample sets with varying concentrations of fat and protein were measured in the M...To compare mid-infrared(MIR)and near-infrared(NIR)spectroscopies for the determination of the fat and protein contents in milk,the same sample sets with varying concentrations of fat and protein were measured in the MIR range of 3 200-700 cm-1 and NIR range of 9 000-4 000 cm-1.The spectral features in the two regions were analyzed.The MIR spectra of milk were characteristic due to the MIR inherent molecular specificity,whereas the NIR spectra were relatively characterless due to the NIR low selectivity.Partial least squares(PLS)regression models for fat and protein were developed by using both MIR and NIR spectra.MIR data with no pretreatment gave better results than NIR data.The square correlation coefficient(R2)and the root mean square error of prediction(RMSEP)were 0.98 and 0.10 g/dL for fat and 0.97 and 0.11 g/dL for protein.With NIR techniques,satisfactory results were not obtained with raw data.However,NIR data after pretreatment gave similarly good results to the ones using MIR method.This paper indicates that either of the MIR and NIR spectral methods is reliable for the determination of the fat and protein contents.展开更多
Objective To investigate the main proteinases responsible for CD16b shedding under different stimulators. Methods HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then ...Objective To investigate the main proteinases responsible for CD16b shedding under different stimulators. Methods HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then overexpressed with a disintegrin and metalloproteinase 10 (ADAM10) or ADAM17, sup- pressed with short hairpin RNA of ADAM10 or ADAM I 7, and reconstituted with ADAM 10 or ADAM17, respectively. After each treatment, the cell line was stimulated with ionomycin or phorbol 12-myristate- 13-acetate (PMA) for 12 hours. The soluble CD 16b released from cell membrane was detected by immuno- precipition and immunoblot. Quantitation was then implemented to compare the amount of soluble CD 16b in cell supernatant after stimulation. Results HEK293 cell line stably expressing CD16b was successfully established. When CDI6b ex- pressing cell line was overexpressed with ADAM 10, shedding of CD 16b was increased after stimulation with ionomycin but not PMA; when the cell line overexpressed with ADAM I7, shedding of CDI6b was increased after stimulation with PMA but not ionomycin. Similarly, when ADAM10 was suppressed by short hairpin RNA, CD 16b shedding was decreased after stimulation with ionomycin; when ADAM 17 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with PMA. The shedding of CD16b was increased again when CD16b expressing cell line was reconstituted with ADAM10 and stimulated by ionomycin or reconstituted with ADAM 17 and stimulated by PMA. Conclusions Both ADAM10 and ADAM17 could shed CD16b, but they possess differed prefer- ences. ADAM10 is the main sheddase under stimulation of ionomycin, while ADAM17 is the main sheddase under stimulation of PMA.展开更多
Distributed polarization coupling in polarization-maintaining fibers can be detected by using a white light Michelson interferometer.This technique usually requires that only one polarization mode is excited.However,i...Distributed polarization coupling in polarization-maintaining fibers can be detected by using a white light Michelson interferometer.This technique usually requires that only one polarization mode is excited.However,in practical measurement,the injection polarization direction could not be exactly aligned to one of the principal axes of the PMF,so the influence of the polarization extinction ratio should be considered.Based on the polarization coupling theory,the influence of the incident polarization extinction on the measurement result is evaluated and analyzed,and a method for distributed polariza-tion coupling detection is developed when both two orthogonal eigenmodes are excited.展开更多
DNA methylation, catalyzed by DNA methyltransferases(MTases), is a key component of genetic regulation, and DNA MTases have been regarded as potential targets in anticancer therapy. Herein, based on our previously dev...DNA methylation, catalyzed by DNA methyltransferases(MTases), is a key component of genetic regulation, and DNA MTases have been regarded as potential targets in anticancer therapy. Herein, based on our previously developed DNA-mediated supercharged green fluorescent protein(Sc GFP)/graphene oxide(GO) interaction, coupled with methylation-initiated template-free DNA polymerization, we propose a novel fluorescence assay strategy for sensitive detection of DNA MTase activity. A hairpin DNA with a methylation-sensitive site and an amino-modified 3′-terminal(DNA-1) was designed and worked as a starting molecule. In the presence of DNA MTase, methylation-sensitive restriction endonuclease, and terminal deoxynucleotidyl transferase(Td T), DNA-1 can be sequentially methylated, cleaved, and further elongated. The resulting long DNA fragments quickly bind with Sc GFP and form the Sc GFP/DNA nanocomplex. Such nanocomplex can effectively protect Sc GFP from being adsorbed and quenched by GO. Without the methylation-initiated DNA polymerization, the fluorescence of Sc GFP will be quenched by GO. Thus, the DNA MTase activity, which is proportional to the amount of DNA polymerization products, can be measured by reading the fluorescence of Sc GFP/GO. The method was successfully used to detect the activity of DNA adenine methylation(Dam) MTase with a wide linear range(0.1–100 U/m L) and a low detection limit of 0.1 U/m L. In addition, the method showed high selectivity and the potential to be applied in a complex sample. Furthermore, this study was successfully extended to evaluate the inhibition effect of 5-fluorouracil on Dam MTase activity and detect Td T activity.展开更多
A sulfonated 9,10-distyrylanthracene derivative with aggregation-induced emission (AIE) property is designed and synthesized. It shows a highly sensitive and selective fluorescence enhancement property for bovine seru...A sulfonated 9,10-distyrylanthracene derivative with aggregation-induced emission (AIE) property is designed and synthesized. It shows a highly sensitive and selective fluorescence enhancement property for bovine serum albumin (BSA) protein detection and quantification. Analysis on the interaction between the probe molecule and BSA reveals the essential role of the hydrophobic cavities of the protein folding structure.展开更多
Quantitative analysis of interactions between small molecules and proteins is a central challenge in chemical genetics, molecular diagnostics and drug developments. Here, we developed a RNA transcription nanomachine b...Quantitative analysis of interactions between small molecules and proteins is a central challenge in chemical genetics, molecular diagnostics and drug developments. Here, we developed a RNA transcription nanomachine by assembling T7 RNA polymerase on a small molecule-labeled DNA heteroduplex. The nanomachine, of which the RNA transcription activity can be quantitatively inhibited by protein binding, showed a great potential for small molecule-protein interaction assay. This finding enabled us to develop a novel homogeneous label-free strategy for assays of interactions between small molecules and their protein receptors. Three small molecule compounds and their protein receptors have been used to demonstrate the developed strategy. The results revealed that the protein-small molecule interaction assay strategy shows dynamic responses in the concentration range from 0.5 to 64 nM with a detection limit of 0.2 nM. Due to its label-free, homogeneous, and fluorescence-based detection format, besides its desirable sensitivity this technique could be greatly robust, cost-efficient and readily automated, implying that the developed small molecule-protein interaction assay strategy might create a new methodology for developing intrinsically robust, sensitive and selective platforms for homogeneous protein detection.展开更多
Visible and infrared(VIR) measurements and the retrieved cloud parameters are commonly used in precipitation identification algorithms, since the VIR observations from satellites, especially geostationary satellites, ...Visible and infrared(VIR) measurements and the retrieved cloud parameters are commonly used in precipitation identification algorithms, since the VIR observations from satellites, especially geostationary satellites, have high spatial and temporal resolutions. Combined measurements from visible/infrared scanner(VIRS) and precipitation radar(PR) aboard the Tropical Rainfall Measuring Mission(TRMM) satellite are analyzed, and three cloud parameters, i.e., cloud optical thickness(COT), effective radius(Re), and brightness temperature of VIRS channel 4(BT4), are particularly considered to characterize the cloud status. By associating the information from VIRS-derived cloud parameters with those from precipitation detected by PR, we propose a new method for discriminating precipitation in daytime called Precipitation Identification Scheme from Cloud Parameters information(PISCP). It is essentially a lookup table(LUT) approach that is deduced from the optimal equitable threat score(ETS) statistics within 3-dimensional space of the chosen cloud parameters. South and East China is selected as a typical area representing land surface, and the East China Sea and Yellow Sea is selected as typical oceanic area to assess the performance of the new scheme. It is proved that PISCP performs well in discriminating precipitation over both land and oceanic areas. Especially, over ocean, precipitating clouds(PCs) and non-precipitating clouds(N-PCs) are well distinguished by PISCP, with the probability of detection(POD) near 0.80, the probability of false detection(POFD) about 0.07, and the ETS higher than 0.43. The overall spatial distribution of PCs fraction estimated by PISCP is consistent with that by PR, implying that the precipitation data produced by PISCP have great potentials in relevant applications where radar data are unavailable.展开更多
文摘AIM: To detect the new serum biomarkers for colorectal cancer (CRC) by serum protein profiling with surfaceenhanced laser desorption ionisation - time of flight mass spectrometry (SELDI-TOF MS). METHODS: Two independent serum sample sets were analysed separately with the ProteinChip technology (set A: 40 CRC + 49 healthy controls; set B: 37 CRC + 31 healthy controls), using chips with a weak cation exchange moiety and buffer pH 5. Discriminative power of differentially expressed proteins was assessed with a classification tree algorithm. Sensitivities and specificities of the generated classification trees were obtained by blindly applying data from set A to the generated trees from set B and vice versa. CRC serum protein profiles were also compared with those from breast, ovarian, prostate, and non-small cell lung cancer. RESULTS: Mass-to-charge ratios (m/z) 3.1×10^3, 3.3× 10^3, 4.5×10^3, 6.6×10^3 and 28×10^3 were used as classitiers in the best-performing classification trees. Tree sensitivities and specificities were between 65% and 90%.Host of these discriminative m/z values were also different in the other tumour types investigated. M/z 3.3× 10^3, main classifier in most trees, was a doubly charged form of the 6.6× 10^3-Da protein. The latter was identified as apolipoprotein C-I. M/z 3.1×10^3 was identified as an N-terminal fragment of albumin, and m/z 28× 10^3 as apolipoprotein A-I. CONCLUSION: SELDI-TOF MS followed by classification tree pattern analysis is a suitable technique for finding new serum markers for CRC. Biomarkers can be identified and reproducibly detected in independent sample sets with high sensitivities and specificities. Although not specific for CRC, these biomarkers have a potential role in disease and treatment monitoring.
文摘Objective: To establish a sensitive and specific system for genotoxicity detection in vivo. Methods: Endogenous p53 tracer vector p53RE was constructed by using green fluorescent protein (GFP) as a reporter to trace p53 transcriptional activity under the control of base SV40 early promoter. The tracer cells 3T3-REG were established by transfecting NIH3T3 cells with tracer vector and treated with ultraviolet, H2O2 and adriamycin to induce DNA damage and the subsequent endogenous p53 expression. The GFP expression and its green fluorescence in the tracer cells were observed and measured with fluorescent microscope and FACS. Results: The GFP expression increased rapidly after various treatment and reached the maximum 1 h later, and decreased gradually afterwards. FACS analysis showed that GFP expression in 3T3-REG tracer cells was consistent with the concentration of H2O2, while that in 3T3-SVG cells, which were transfected with control vector pSV-GFP. hardly increased in response to the treatment. Conclusion: GFP tracing p53 transcriptional activity is a sensitive and specific method for genotoxicity detection.
基金Supported by National Natural Science Foundation of China(No.30170261)the 10th Five-Year Plan of China(No.2004BA706B12).
文摘To compare mid-infrared(MIR)and near-infrared(NIR)spectroscopies for the determination of the fat and protein contents in milk,the same sample sets with varying concentrations of fat and protein were measured in the MIR range of 3 200-700 cm-1 and NIR range of 9 000-4 000 cm-1.The spectral features in the two regions were analyzed.The MIR spectra of milk were characteristic due to the MIR inherent molecular specificity,whereas the NIR spectra were relatively characterless due to the NIR low selectivity.Partial least squares(PLS)regression models for fat and protein were developed by using both MIR and NIR spectra.MIR data with no pretreatment gave better results than NIR data.The square correlation coefficient(R2)and the root mean square error of prediction(RMSEP)were 0.98 and 0.10 g/dL for fat and 0.97 and 0.11 g/dL for protein.With NIR techniques,satisfactory results were not obtained with raw data.However,NIR data after pretreatment gave similarly good results to the ones using MIR method.This paper indicates that either of the MIR and NIR spectral methods is reliable for the determination of the fat and protein contents.
基金Supported by the National Natural Science Foundation of China (30872287)
文摘Objective To investigate the main proteinases responsible for CD16b shedding under different stimulators. Methods HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then overexpressed with a disintegrin and metalloproteinase 10 (ADAM10) or ADAM17, sup- pressed with short hairpin RNA of ADAM10 or ADAM I 7, and reconstituted with ADAM 10 or ADAM17, respectively. After each treatment, the cell line was stimulated with ionomycin or phorbol 12-myristate- 13-acetate (PMA) for 12 hours. The soluble CD 16b released from cell membrane was detected by immuno- precipition and immunoblot. Quantitation was then implemented to compare the amount of soluble CD 16b in cell supernatant after stimulation. Results HEK293 cell line stably expressing CD16b was successfully established. When CDI6b ex- pressing cell line was overexpressed with ADAM 10, shedding of CD 16b was increased after stimulation with ionomycin but not PMA; when the cell line overexpressed with ADAM I7, shedding of CDI6b was increased after stimulation with PMA but not ionomycin. Similarly, when ADAM10 was suppressed by short hairpin RNA, CD 16b shedding was decreased after stimulation with ionomycin; when ADAM 17 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with PMA. The shedding of CD16b was increased again when CD16b expressing cell line was reconstituted with ADAM10 and stimulated by ionomycin or reconstituted with ADAM 17 and stimulated by PMA. Conclusions Both ADAM10 and ADAM17 could shed CD16b, but they possess differed prefer- ences. ADAM10 is the main sheddase under stimulation of ionomycin, while ADAM17 is the main sheddase under stimulation of PMA.
基金National Natural Science Foundation of China (60577013)the Teaching and Research Award Program for Outstanding Young Teachers of MOE, China
文摘Distributed polarization coupling in polarization-maintaining fibers can be detected by using a white light Michelson interferometer.This technique usually requires that only one polarization mode is excited.However,in practical measurement,the injection polarization direction could not be exactly aligned to one of the principal axes of the PMF,so the influence of the polarization extinction ratio should be considered.Based on the polarization coupling theory,the influence of the incident polarization extinction on the measurement result is evaluated and analyzed,and a method for distributed polariza-tion coupling detection is developed when both two orthogonal eigenmodes are excited.
基金supported by the National Basic Research Program (2011CB911002)the National Natural Science Foundation of China (21190044, 21475037, 21222507, 21175036)the fundamental research funds for the central universities
文摘DNA methylation, catalyzed by DNA methyltransferases(MTases), is a key component of genetic regulation, and DNA MTases have been regarded as potential targets in anticancer therapy. Herein, based on our previously developed DNA-mediated supercharged green fluorescent protein(Sc GFP)/graphene oxide(GO) interaction, coupled with methylation-initiated template-free DNA polymerization, we propose a novel fluorescence assay strategy for sensitive detection of DNA MTase activity. A hairpin DNA with a methylation-sensitive site and an amino-modified 3′-terminal(DNA-1) was designed and worked as a starting molecule. In the presence of DNA MTase, methylation-sensitive restriction endonuclease, and terminal deoxynucleotidyl transferase(Td T), DNA-1 can be sequentially methylated, cleaved, and further elongated. The resulting long DNA fragments quickly bind with Sc GFP and form the Sc GFP/DNA nanocomplex. Such nanocomplex can effectively protect Sc GFP from being adsorbed and quenched by GO. Without the methylation-initiated DNA polymerization, the fluorescence of Sc GFP will be quenched by GO. Thus, the DNA MTase activity, which is proportional to the amount of DNA polymerization products, can be measured by reading the fluorescence of Sc GFP/GO. The method was successfully used to detect the activity of DNA adenine methylation(Dam) MTase with a wide linear range(0.1–100 U/m L) and a low detection limit of 0.1 U/m L. In addition, the method showed high selectivity and the potential to be applied in a complex sample. Furthermore, this study was successfully extended to evaluate the inhibition effect of 5-fluorouracil on Dam MTase activity and detect Td T activity.
基金the National Basic Research Pro- gram of China (973 Program, 2013CB834702, 2009CB623605)the Natural Science Foundation of China (21074045, 21204027, 21221063)+1 种基金the Research Fund for the Doctoral Program of Higher Education of China (20120061120016)the Project of Jilin Province (20100704)
文摘A sulfonated 9,10-distyrylanthracene derivative with aggregation-induced emission (AIE) property is designed and synthesized. It shows a highly sensitive and selective fluorescence enhancement property for bovine serum albumin (BSA) protein detection and quantification. Analysis on the interaction between the probe molecule and BSA reveals the essential role of the hydrophobic cavities of the protein folding structure.
基金supported by the National Natural Science Foundation of China (21025521, 21035001&20875027)the National Key Basic Re-search Program (2011CB911000)+3 种基金European Commission FP7-HEALTH-2010 Programme-GlycoHIT (260600)National Grand Program on Key Infectious Disease (2009ZX10004-312)Postdoctoral Science Foundation (20100480934) of ChinaChangjiang Scholars and Innovative Research Team in University Program and Natural Science Foundation of Hunan Province (10JJ7002)
文摘Quantitative analysis of interactions between small molecules and proteins is a central challenge in chemical genetics, molecular diagnostics and drug developments. Here, we developed a RNA transcription nanomachine by assembling T7 RNA polymerase on a small molecule-labeled DNA heteroduplex. The nanomachine, of which the RNA transcription activity can be quantitatively inhibited by protein binding, showed a great potential for small molecule-protein interaction assay. This finding enabled us to develop a novel homogeneous label-free strategy for assays of interactions between small molecules and their protein receptors. Three small molecule compounds and their protein receptors have been used to demonstrate the developed strategy. The results revealed that the protein-small molecule interaction assay strategy shows dynamic responses in the concentration range from 0.5 to 64 nM with a detection limit of 0.2 nM. Due to its label-free, homogeneous, and fluorescence-based detection format, besides its desirable sensitivity this technique could be greatly robust, cost-efficient and readily automated, implying that the developed small molecule-protein interaction assay strategy might create a new methodology for developing intrinsically robust, sensitive and selective platforms for homogeneous protein detection.
基金supported by the National Basic Research Program of China (Grant No. 2010CB428601)the Strategic Priority Research Program-Climate Change (Carbon Budget and Relevant Issues of the Chinese Academy of Sciences) (Grant No. XDA05100303)+2 种基金the Fundamental Research Funds for the Central Universities (Grant No. WK2080000024)the National Natural Science Foundation of China (Grant Nos. 41230419, 41175032 and 41075041)the Guangdong Science and Technology Plan Project (2012A061400012, 2011A032100006)
文摘Visible and infrared(VIR) measurements and the retrieved cloud parameters are commonly used in precipitation identification algorithms, since the VIR observations from satellites, especially geostationary satellites, have high spatial and temporal resolutions. Combined measurements from visible/infrared scanner(VIRS) and precipitation radar(PR) aboard the Tropical Rainfall Measuring Mission(TRMM) satellite are analyzed, and three cloud parameters, i.e., cloud optical thickness(COT), effective radius(Re), and brightness temperature of VIRS channel 4(BT4), are particularly considered to characterize the cloud status. By associating the information from VIRS-derived cloud parameters with those from precipitation detected by PR, we propose a new method for discriminating precipitation in daytime called Precipitation Identification Scheme from Cloud Parameters information(PISCP). It is essentially a lookup table(LUT) approach that is deduced from the optimal equitable threat score(ETS) statistics within 3-dimensional space of the chosen cloud parameters. South and East China is selected as a typical area representing land surface, and the East China Sea and Yellow Sea is selected as typical oceanic area to assess the performance of the new scheme. It is proved that PISCP performs well in discriminating precipitation over both land and oceanic areas. Especially, over ocean, precipitating clouds(PCs) and non-precipitating clouds(N-PCs) are well distinguished by PISCP, with the probability of detection(POD) near 0.80, the probability of false detection(POFD) about 0.07, and the ETS higher than 0.43. The overall spatial distribution of PCs fraction estimated by PISCP is consistent with that by PR, implying that the precipitation data produced by PISCP have great potentials in relevant applications where radar data are unavailable.