Nowadays, proteomics focuses on quantitative analysis rather than qualitative. In the field of quantitative proteomics, Isobaric tags for relative and absolute quantitation (iTRAQ) is one of the most widely used techniques. The advantage of iTRAQ is high throughput, high stability and free of the restriction of sample property. iTRAQ is suitable for almost all kinds of samples, and up to 8 samples can be analyzed simultaneously by commercially available kit. Along with the development of techniques, more and more mass spectrometry (MS) platforms are used in iTRAQ experiments and the accuracy of iTRAQ has been improved. iTRAQ has been applied to studies of microorganism, animal, plant, medical and protein post-translational modification. Here we review the recent progress in the development of iTRAQ and its applications in quantitative proteomics.
Animals ; Humans ; Mass Spectrometry ; Proteomics ; methods

This paper developed a calculation method to acquire the yield load P of bone plate during four-point bending test. This method is based on the displacement--force (δ-F) curve function f(M)(δ) obtained from the test, each slope of the curve was calculated using piecewise smooth function and the line segment in f(M)(δ) elastic deformation area was searched by setting the minimum slope T. Slope S was obtained through linear fit so as to build parallel displacement function f(L)(δ). Then, approximating intersection point of f(M)(δ) and f(L)(δ) was obtained through linear interpolation. Thus, yield load P was acquired. The method in the paper was loyal to YY/T 0342-2002 regulation and was liable to program calculation. The calculating process was nothing to do with whether the initial point during the test was preloaded or unloaded, and there was no need to correct the original point. In addition, T was set in an ideal fitting level guaranteed by the fitting coefficient of determination R2, and thus S was very close to the real value, and P was with a high accuracy.
Biomechanical Phenomena ; Bone Plates ; Humans ; Models, Theoretical ; Posture ; Weight-Bearing

A new method based on adherence of cellulolytic bacteria to insoluble cellulose for isolation and purification of thermophilic cellulolytic anaerobes was reported, in which Hungate anaerobic operating techniques were used to roll tubes with insoluble cellulose powder as substrate.

In proteomic research, to improve protein solubility of membrane proteins and nuclear proteins, buffers containing high concentration of detergent, such as 4% SDS, were widely used. However, high concentration of detergent might severely interfere with the downstream proteomic analysis, including protein quantitation and trypsin digestion. To improve the proteomic compatibility of buffers with high concentration of detergent, we used short gel method to pretreat buffers containing detergent. Protein samples were first separated by a short (2-2.5 mm) SDS-PAGE electrophoresis, and proteins were quantitated by comparing with bovine serum albumin standards via optical density analysis. The gel was then cut and peptides were recovered using in-gel digestion. The quantitative linearity range of this method was 1 to 8 μg. The quantitation was accurate and reproducible. After short gel analysis, recovered peptides generated high mass spectrometry signals. In conclusion, short gel method eliminated the interference of high concentration detergent in the proteomics analysis, and it was suitable for protein samples' pretreatment, and was worth to apply in proteomic research.
Detergents ; chemistry ; Electrophoresis, Polyacrylamide Gel ; methods ; Mass Spectrometry ; Membrane Proteins ; chemistry ; Nuclear Proteins ; chemistry ; Proteins ; chemistry ; Proteomics ; methods ; Trypsin

Objective To explore the correlation of the myeloid antigen expression and clinical characteristics of acute lymphoblastic leukemia (ALL) in children.Methods The clinical data of 77 newly diagnosed ALL patients in Department of Hematology,the People's Hospital of Zhengzhou University from Jan.2010 to Dec.2013 were analyzed.The patients included 53 boys and 24 girls with a median age of 7.73 (2.00-15.00) years old.Based on flow cytometry (FCM) analysis of bone marrow,these patients were divided into 2 groups:one group included 26 patients with positive myeloid antigen expression (MyAg + ALL) and the other group included 51 patients with negative myeloid antigen expressions (MyAg-ALL).The correlation among myeloid antigen expression,clinical features,prednisone experiment,myelogram on the 15th day was analyzed through induction chemotherapy and minimal residual disease (MRD) on the 33rd day,and the rate of disease-free survival (DFS) was compared between the 2 groups.Results There were 26 cases with myeloid antigen expression among 77 patients (33.77％),CD13 + accounting for 19.48％ (15/77 cases),CD33 + 10.39％ (8/77 cases),and CD117 + 5.19％ (4/77 cases).Among these patients,there were 2 patients expressing both CD13 + and CD33 +,and 1 patient expressing both CD33 + and CD117 +.There was no difference between the MyAg + ALL group and MyAg-ALL group in gender (x2 =0.217,P =0.641),age (≥ 10 years old,x2 =0.011,P =0.918),white blood count(≥50 × 109/L,x2 =1.198,P =0.274),lactate dehydrogenase (LDH) (≥500 U/L,x2 =0.317,P =0.573),genetic abnormality (x2 =0.377,P =0.539),immunophenotype (B-ALL/T-ALL,x2 =0.397,P =0.529),and risk stratification (low-risk group,middle-risk group and high-risk group,x2 =0.260,P =0.878).Univariate Logistic regression showed that the reaction rate of prednisone experiment (P =0.023,OR =3.422) and positive rate of MRD (P =0.001,OR =0.133) of MyAg + ALL group were obviously higher than those in MyAg-ALL group.Multivariate Logistic regression showed that positive rate of MRD in CD13 + ALL group was obviously higher than that of CD13-ALL group (P =0.034,OR =120.765).The DFS rate of CD13 + ALL group and CD13-ALL group were (50.4 ± 13.8)％ and (77.4 ±6.7)％ respectively,and there was a significant difference between the 2 groups (x2 =3.928,P =0.047).Conclusions There is no significant correlation between myeloid antigen expression and clinical characteristics of children patients with ALL.For the patients with myeloid antigens,the early reaction of induction chemotherapy is bad,and for patients with CD13,the prognosis is not good.

Objective To evaluate the outcome of the procedure of using the free flap vascularized by the dorsal artery of big toe to treat finger skin defect.Methods From August 2010 to April 2013,21 cases of finger shin defect were treated with the free flap vascularized by the dorsal artery of big toe in which emergency surgeries were conducted in 9 cases and sub emergency surgeries were conducted in 12 cases.The age of the 21 patients was 21 to 48 years old and 15 of them were males and 6 were females.Thumb was involved in 6 patients and index finger was involved in 15 patients.The skin defect occurred at dorsal in 7 patients and palm in 14 patients.The area of the flaps ranged from 2.2 cm × 1.6 cm to 4.0 cm × 3.2 cm.Observed the restoration of the affected fingers' appearance and function,investigated the clinical results and concluded the indication and advantages and disadvantages of this procedure by following up.Results All of the 21 flaps survived at the last office visit.The follow-up period ranged from 3 to 18 months.The shapes and the fingerprints of the flaps were satisfied.The color and texture of the flaps were similar to those of the finger skin.The 2 points discriminations of the flaps was 6-8 mm.No function deficits were found in the donor feet.Conclusion This free flap is satisfied in the shape,easy to harvested and the blood supply was constant in its location,and recommend it in treating the small of middle area skin defect in the finger.

We purified a sarcosine oxidase from Bacillus sp. strain BSD-8 isolated from soil. We purified the enzyme by ammonium sulfate precipitation, DEAE-cellulose, Toyopearl hydrophobic and Sephadex G-75 molecular sieve chromatography and characterized the purified sarcosine oxidase. This sarcosine oxidase was a flavin enzyme containing a noncovalently bound flavin with the subunit molecular mass of 51 kDa. The optimal temperature for this enzyme was 60 degrees C and it showed its highest activity at pH 8.5. It was stable in the pH range of 8.0-10.0 and at the temperature of 60 degrees C. Estimated by Lineveaver-Burk plots, the K(m) of the enzyme was 3.1 mmol/L. Ag+, Hg2+, SDS and Tween 80 dramatically inhibted the enzyme activity, whereas Tween 20 and Triton X-100 had no effect on enzyme activity. The thermostability of this enzyme was better than reported sarcosine oxidases, and it could be applied in enzymatic measuring of creatinine.
Bacillus ; enzymology ; isolation & purification ; Bacterial Proteins ; chemistry ; isolation & purification ; metabolism ; Chemical Precipitation ; Enzyme Stability ; Sarcosine Oxidase ; chemistry ; isolation & purification ; metabolism ; Soil Microbiology

In order to characterize a thermostable urate oxidase (Uox) from Microbacterium sp. strain ZZJ4-1, we cloned its gene (uox). The open reading frame of uox contained 894 base pairs and encoded a protein with 297 amino acids. Alignment of gene sequences indicated there was no obvious identity with the most reported uox and that 72% identity was found with uox from Arthrobacter globiformis. We inserted the gene into the plasmid pET-15b to construct an expression vector pET-15b-uox and got it induced expression in Escherichia coli BL21 (DE3). After the purification of the recombinant Uox by the HisBind column, we studied some properties of it. It was composed of subunits with a molecular mass of about 35 kDa. The optimal temperature and pH was 30 degrees C and pH 7.5. It was stable below 65 degrees C and from pH 8.5 to 11.0. The Km value was 0.22 mmol/L with the uric acid as the substrate. Ag+, Zn2+, CU2+ and SDS could totally inhibit its activity while Tween 20, Tween 80 and Triton X-100 had a slight promotion effect. The thermal stability of this enzyme was the most excellent among the reported recombinant Uox. Based on this property, it would be very useful in the application.
Actinomycetales ; enzymology ; genetics ; Amino Acid Sequence ; Cloning, Molecular ; Enzyme Stability ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Molecular Sequence Data ; Recombinant Proteins ; biosynthesis ; genetics ; Urate Oxidase ; genetics ; metabolism

Objective Discuss the clinical efficacy by using two kinds of mini perforator free flap for digital injuries reconstruction. Methods From August, 2014 to February, 2017, 45 patients were managed randomly with either radial artery superficial palmar branch(RASPB)perforator free flap or digital artery(DA)perforator free flap for digital skin defects reconstruction, and they were therefore divided into two groups according to the flap type. There were 24 patients in RASPB group, with an average wound dimensions ranged from 1.8 cm×1.5 cm to 4.0 cm×2.5 cm, and an average harvested flap size ranged from 2.0 cm×1.7 cm to 4.2 cm×2.6 cm. Another 21 patients were in DA group, with an average wound dimensions ranged from 2.0 cm×1.5 cm to 3.8 cm×3.0 cm, and an average harvested flap size ranged from 2.2 cm×1.6 cm to 3.9 cm×3.2 cm. The survival rate, sensory function, donor site complications, hand function recovery and aesthetic outcomes of two groups were compared by the SPSS22.0 statistical software after surgery. Results The mean follow up period was 15 months. All flaps were primary survived without vascular crisis. The flaps were soft in texture,trimness in appearance and none of them overtop the normal skin for more than 0.5 cm. Both groups had a favorable sensory recovery.All cases recovery to S3+or better.In Group RASPB,the mean two point discrimination(2 PD)was 7.85±1.15 mm(ranged from 7.0 mm to 9.0 mm). And it was 6.02±0.94 mm(ranged from 6.0 mm to 8.0 mm)in DA group. The difference between two groups was statistically significant(P <0.05). Then we synthetically analyzed flap texture and sensory function,and calculated the qualified ratio of each group.There was no significant difference between two groups(P > 0.05). The degree of scar contracture demonstrated donor site compli cations in RASPB group was lesser than that in DA group(P<0.05).The range of motion of interphalangeal joint was used to reflect the hand function. And we calculated the ratio of repaired and contralateral sites. The difference of the mean ratio between two groups was not statistically significant(P>0.05). Conclusion On account of the characteristics of invariant anatomy position, sufficient blood supply, favorable aesthetic outcome and minimal donor site mobility, both RASPB perforator flap and DA perforator flap were optimal for digital skin defects reconstruction.Besides,incorporated with nerve and tendon,the RASPB perforator flap can also be used for complex tissue transplantation,and the surgery field was only on the arm.While the DA perforator free flap had an advantage of better sensory recovery and appearance.

Coronaviruses are the causative agent of respiratory and enteric diseases in animals and humans. One example is SARS, which caused a worldwide health threat in 2003. In coronaviruses, the structural protein N (nucleocapsid protein) associates with the viral RNA to form the filamentous nucleocapsid and plays a crucial role in genome replication and transcription. The structure of N-terminal domain of MHV N protein also implicated its specific affinity with transcriptional regulatory sequence (TRS) RNA. Here we report the crystal structures of the two proteolytically resistant N- (NTD) and C-terminal (CTD) domains of the N protein from murine hepatitis virus (MHV). The structure of NTD in two different crystal forms was solved to 1.5 Å. The higher resolution provides more detailed structural information than previous reports, showing that the NTD structure from MHV shares a similar overall and topology structure with that of SARS-CoV and IBV, but varies in its potential surface, which indicates a possible difference in RNA-binding module. The structure of CTD was solved to 2.0-Å resolution and revealed a tightly intertwined dimer. This is consistent with analytical ultracentrifugation experiments, suggesting a dimeric assembly of the N protein. The similarity between the structures of these two domains from SARS-CoV, IBV and MHV corroborates a conserved mechanism of nucleocapsid formation for coronaviruses.
Amino Acid Sequence ; Binding Sites ; Crystallography, X-Ray ; Molecular Sequence Data ; Murine hepatitis virus ; chemistry ; metabolism ; Nucleocapsid Proteins ; chemistry ; metabolism ; Phosphoproteins ; chemistry ; metabolism ; Protein Binding ; Protein Folding ; Protein Multimerization ; Protein Structure, Secondary ; Protein Structure, Tertiary ; RNA ; metabolism ; Sequence Alignment