To construct a library of mouse single chain variable fragment (scFv) antibody against ofloxacin using phage display and recombinant antibody technique, specific anti-ofloxacin scFv was screened and 3D structure was homology modeling. Total RNA was extracted from hybridoma cell of ofloxacin mAb, and was used to amplify VH and VL gene by RT-PCR using random primer. Then they were linked by a DNA linker encoding (G1y4 Ser) 3 as VH-linker-VL sequence forming scFv by SOE(splicing by overlap extension) PCR. These fragments were inserted into phage T7 after double digestion and transformed with host bacteria BLT5403. 3 ×105 pfu / ml single chain antibody phage libraries were successfully constructed. Four positive phage scFv clones were screened by direct competitive ELISA after four times of enriched procedure in the order of adsorption-elution-amplificatio, 3D structure of specific scFv was homology modeling finally. This research lays a foundation for further massive expression of anti-ofloxacin scFv.

The method of flying through the air is a qi-promoting and qi-circulating technique commonly used in clinical acupuncture. It includes four methods: the blue dragon wagging its tail, the white tiger shaking its head, the green turtle probing the cave and the red phoenix winging to the source and functions to circulate bodily meridian qi. The method of flying through the air was firstrecorded in Golden needle Fu. Later and modern doctors developed it on the basis of Golden needle Fu. This article straightens up the historical origin and development of four methods of flying through the air.

Objective To investigate the feasibility of modified rapid Carba NP test for the detection of carbapenemase,and analyze the differences between the modified method and Carba NP test.Methods A total of 264 strains of gram-negative bacillus,including 164 carbapenem-resistant strains and 100 sensitive strains,were collected,and their carbapenemase were detected by Carba NP test and the modified rapid Carba NP test,respectively.The differences between the two tests were evaluated based on PCR as a reference.Results Among 164 carbapenem-resistant strains,carbapenemase gene was detected in 144 strains by PCR.The carbapenemase gene was negative in 100 sensitive strains.Among 164 carbapenem-resistant strains,135 were positive for the Carba NP test,while 130 for the modified rapid Carba NP test.One hundred of sensitive strains were negative for the two Carba NP tests.Compared with the results of PCR,the sensitivity,specificity and Kappa value of the Carba NP test were 91.7％ (132/144),97.5％ (117/120) and 0.886,respectively,while those of the modified rapid Carba NP test were 89.6％ (129/144),99.2％ (119/120) and 0.879,respectively.There was no significant difference in the positive rates between Carba NP test and the modified rapid Carba NP test (x2 =1.45,P ＞ 0.05).Conclusion The modified rapid Carba NP test which has high consistency with the PCR method,is faster and cheaper than the Carba NP test,and may be applied to epidemiologic survey and the early monitoring of nosocomial infections.

Objective To evaluate the application value of inhibitor enhanced modified carbapenemase inactivation method (imCIM) in the detection of class B carbapenemase.The differences between imCIM and EDTA disc potentiation test (EDPT) were comparatively analyzed.Methods A total of 181 strains of carbapenem insensitive strains were collected,among which there were 44 strains of Klebsiella pneumoniae,44 strains of Escherichia coli,43 strains of Acinetobacter baumannii and 50 strains of Pseudomonas aeruginosa.The 83 strains of carbapenem-sensitive strains were composed of 25 strains of Klebsiella pneumoniae,16 strains of Escherichia coli,25 strains of Acinetobacter baumannii and 17 strains of Pseudomonas aeruginosa.The class B carbapenemase in the 264 strains of pathogenic bacteria was screened by imCIM and EDPT,and PCR results were used as gold standard.The statistical analysis wasperformed with consistency check,related-sample Wilcoxon signed rank sum test,independent samples Kruskal-Wallis H test and ROC curve.Results Among the 181 strains of carbapenem insensitive strains,PCR results of 144 strains were positive for drug resistance gene.The samples of class A,B and D of carbapenemase were 39,77 and 28 strains respectively.The results of imCIM showed that 70 strains were positive,and the other 111 strains were negative.The imCIM results of 166 strains were consistent with those of PCR.The results of EDPT showed that 72 strains were positive,and the other 109 strains were negative.The EDPT results of 134 strains were consistent with those of PCR.The results of PCR,EDPT and imCIM of 83 carbapenem sensitive strains were negative.The sensitivity and specificity of imCIM were 85.71％ (66/77) and 97.86％ (183/187),and the value of Kappa was 0.859.The sensitivity and specificity of EDPT were 66.23 ％ (51/77) and 88.77 ％ (166/187),and the value of Kappa was 0.561.The difference of inhibition zone of imCIM (AdimCIM) was different from EDPT(AdEDPr) and the difference was statistically significant (Z =-6.941,P ＜ 0.05).In the imCIM detection,the AdimciM level of class B carbapenemase showed different population distribution position from class A and D carbapenemase with the statistically significant difference (x2 =108.887,P ＜ 0.05).The areas under the ROC curve of imCIM and EDPTwere 0.988 (95％CI:0.977 to0.999) and0.936 (95％CI:0.909 to0.963),respectively.Conclusion imCIM should be accurate,efficient and convenient for screening of carbapenem phenotype for its high sensitivity and specificity,and suitable for epidemiological monitoring.

OBJECTIVETo discuss the replantation of fingertip amputation in lack of availability of intravenous anastomosis.

METHODSFrom November 2009 to November 2010, 86 patients (104 fingers) with fingertip amputation were treated with replantatioin, including 64 males and 22 females, with an average age of 26 years ranging from 2 to 64 years. The time from injury to therapy was from 30 min to 12 h, time of broken finger ischemia was from 2.5 to 12 h. Preoperative examination showed no obvious abnormalities. Four different replantation methods were selectively applied to these 104 amputated fingertips of 86 cases: (1) replantation with anastomosis of single or bilateral proper digital artery in 37 fingers; (2) replantation with arteriovenous bypass in 27 fingers; (3) replantation with exclusive anastomosis of digital artery in 24 fingers; (4) replantation with removing the palmar pocket method in 16 fingers.

RESULTSOne hundred and two of 104 amputated fingertips were survived. Among these survived fingers,75 cases (92 fingers) were followed-up for 6 to 24 months. According to the assessment standard of Chinese Medical Association of Hand Surgery, the results were excellent in 52 cases, good in 19, poor in 4.

CONCLUSIONIt benefits to expand the indications and improve the survival rate of replantation of fingertip amputation with the correct choice of different replantation methods according to the injury situation of the broken fingertip artery after debridement under the microscope.

Adolescent ; Adult ; Amputation ; Child ; Child, Preschool ; Debridement ; Female ; Fingers ; physiology ; surgery ; Humans ; Male ; Middle Aged ; Recovery of Function ; Replantation ; methods ; Young Adult

Objective To evaluate the feasibility and effectiveness of four closure techniques ,large incision with plastic distal at‐tachment and clip with suction ,1 .1 cm small incision with plastic distal attachment and clip with suction in natural orifice translu‐minal endoscopic surgery(NOTES) .Methods Forty‐one ex vivo porcine stomachs were involved in this research .According to the size of incision and different methods of incision closure ,all ex vivo porcine stomachs were divided into four groups .Group A in‐volved 8 ex vivo porcine stomachs ,their size of incision were 1 .2-2 .0 cm ,and their incisions were closured by a clip closure direct‐ly ;group B involved 10 ex vivo porcine stomachs ,their size of incision were 1 .2-2 .0 cm ,and their incisions were closed by plastic end attachment with suction and clip enclosure technique;group C involved 10 ex vivo porcine stomachs ,their size of incision were smaller than 1 .1 cm ,and their incisions were by a clip closure directly ;group D involved 13 ex vivo porcine stomachs ,their size of incision were smaller than 1 .1 cm ,and closured by plastic end attachment with suction and clip enclosure technique .Stomach leaks were evaluated by leaking studies after the procedure .Results In group A ,1 incision was closed successfully ,7 incisions were clo‐sured unsuccessfully ;in group B ,2 incisions were closed successfully ,8 incisions were closed unsuccessfully ;in group C ,2 incisions were closed successfully ,8 incisions were closured unsuccessfully ;in group D ,11 incisions were closed successfully ,2 incisions were Closured unsuccessfully .Fisher′s exact test showed that group D was significantly related to the success of incision closure .The were significant differences between group A and D ,group B and group D(P<0 .05) .Conclusion Small incision(smaller than 1 cm) and plastic end attachment with suction and clip enclosure technique are optimal in NOTES procedure in this ex vivo porcine stomachs study .

Objective To introduce the operation skills of tail vein injection in conscious rats , and improve the success rate of ingection .Methods The rat was fixed by the operator with a self-made binding clothes , one person assists to fix the rat tail, one person performs puncture , and one person performs injection .During the injection process , we should minimize the injury to the rat tail and the stress caused by operation , strictly limit the needle point , the number of puncture , the maximum dose and injection speed , and to make the needle position away from the injection site to avoid contamination and waste of the drug solution .Results The operation method was successfully established and it was fast , stable, with good repeatability and high degree of coordination .Conclusion This operation is rapid, reliable and stable, worthy of recommendation , especially for the intravenous injection of expensive drugs .

OBJECTIVEReview and discuss anesthesia and operative approach of resection of the trachea for different tracheal diseases, especially for the resection of long-segment of trachea. At the same time to introduce the method of reconstruction of long-segment of trachea with Zhao's (two-stage procedure with memory-alloy mesh) artificial trachea.

METHODSRetrospective study of 18 cases of tracheal resection, analysis of the relation between the choice of anesthetic and operative approach.

RESULTSGeneral anesthesia through cut open the trachea with local anesthesia in 2 cases, general anesthesia through previous tracheotomy in 2 cases, extracorporeal circulation in 2 cases, general anesthesia through endotracheal tube in 12 cases. There were no anesthetic or operative death. Local resection in 3 cases, segmental resection in 15 cases. The longest segmental resection was 8.0 cm. Primary anastomosis after segmental resection in 8 cases, Reconstruction with Zhao's artificial trachea in 7 cases. Postoperative follow-up was 5 months to 8 years. Four cases died from systemic metastasis or other reasons at 4, 11 and 12 months, respectively.

CONCLUSIONSDifferent methods of anesthetic and operative procedures should be used for different patients. Extracorporeal circulation used for patient with highest dangerous condition, or, for which could be inserted endotracheal tube by tracheotomy with local anesthesia. Conservative local resection performed only for patients with very bad general condition. Segmental resection less than 5 cm long could be reconstructed with primary reanastomosis. Resection longer than 5.5 cm could be reconstructed with Zhao's artificial trachea.

Adolescent ; Adult ; Aged ; Anesthesia, General ; methods ; Anesthesia, Local ; Artificial Organs ; Extracorporeal Circulation ; Female ; Humans ; Intubation, Intratracheal ; Male ; Middle Aged ; Prosthesis Implantation ; Retrospective Studies ; Thoracotomy ; Tracheal Diseases ; surgery ; Tracheotomy ; methods

Objective:To investigate the value of procalcitonin (PCT),C-reactive protein (CRP) in discriminating the bloodstream infection bacterial species,and to analyze the levels of PCT and CRP in sepsis caused by different pathogens.Methods:Patients with blood culture and PCT,CRP detection were collected in the study,from January 2015 to January 2017.The levels of PCT and CRP were detected with electrochemiluminescence and immunoturbidimetric,respectively.The levels of PCT in gram positive bacteria and gram negative bacteria,different bacterial species were compared by SPSS21.0 statistical software.Receiver operating characteristic(ROC) curve was established by Sigma software,calculating the optimal cutoff value.Results:The difference in the levels of PCT between gram positive bacteria and gram negative bacteria was statistically significant,and the value of U was 4 420.00,(P=0.004).ROC analysis showed the optimal cut-off value was 1.105 ng/ml.The levels of PCT had statistically significant difference in coagulase-positive staphylococci and coagulase-negative staphylococci,and the value of U was 79.00(P<0.001).ROC analysis revealed the optimal cut-off value of 0.870 ng/ml.There was a significant difference in the levels of PCT in Enterobacteriaceae and non fermentative bacteria and the value of U was 681.50(P=0.005).ROC analysis showed the optimal cutoff value was 3.310 ng/ml.There was no significant difference in PCT between Staphylococcus and Enterococcus.The levels of CRP was no significant difference between the groups.Conclusion:Detection of PCT has certain value in discriminating of bloodstream infection by different bacterial species,which can provide the basis for the early rational use of drugs in patients with suspected bacteremia.The detection of PCT combined with the blood culture,can reduce the risk of failure in patients with severe infection,and improve the efficiency of treatment.

Objective The optimal access for natural orifice transluminal endoscopic surgery is still uncertain .This study was designed to compare the practicability and maneuverability of transgastric ,transunmbilical ,and transrectal approach in abdominal surgery in a canine model .Methods Three dogs were used in this research .Three approach :trangastric ,transunmbilical and tran-srectal approach were carried out for abdominal exploration ,liver biopsy ,bladder biopsy and an attempted cholecystectomy .The ma-neuverability ,endoscopic image ,performer′s perception ,and spatial orientation were evaluated .Results The maneuverability of trangastric ,and transrectal approach NOTES were better than transunmbilical NOTES .Abdominal exploration ,live biopsy ,and bladder biopsy were completed successfully .The cholecystectomy was failed because of poor exposure and difficulty of separating the around tissure .Conclusion The optimal approach for upper abdomen NOTES is transrectal route .For lower abdomen NOTES , the trangastric approach is superior to other accesses .Further study is needed to develop more flexible and precise equipment for NOTES and to evaluate more feasible access approach .