Here we discuss whether N terminal sequencing is appropriate as one of the conventional control methods for monoclonal antibody products. We determined the N terminal sequences of two monoclonal antibody products targeting two antigens separately with both Edman degradation and mass peptide spectrometry. We also identified the characteristic peptide fragments with mass spectrometry. Furthermore, we analyzed their heterogeneity with ion exchange chromatography, capillary zone electrophoresis and Imaged Capillary Isoelectric Focusing. Edman degradation method showed that the N terminal 15 amino acids of heavy and light chains of the two monoclonal antibodies were identical. Peptide mass spectrometry demonstrated that T1 peptide fragments of heavy and light chains of the two antibodies were also the same. But in contrast, peptide mapping and the three analytical methods for heterogeneity analysis could effectively identify and differentiate the two antibodies. The N terminal sequences of two monoclonal antibodies are identical because the number of framework sequences of humanized or human monoclonal antibodies is relatively limited, so whether N terminal sequencing analysis could be regulated as one of the practical control methods should be carefully discussed. Our work also proves that the above analytical methods could combinatorially applied to the identification of monoclonal antibody products, and are more objective compared to N terminal sequencing.
Amino Acid Sequence ; Antibodies, Monoclonal ; isolation & purification ; Chromatography, Ion Exchange ; Humans ; Isoelectric Focusing ; Mass Spectrometry ; Peptide Mapping ; Peptides ; Sequence Analysis, Protein ; methods

Objective To determine the best endoscopic treatment for acute obstructive suppurative cholangitis (AOSC).Methods Data of 93 patients who were diagnosed as having AOSC in endoscopic center in the last three years were retrospectively analyzed.All patients were divided into three groups according to the different treatment methods:ENBD group,ENBD+ERBD group and double ERBD group.Postoperative temperature,bilirubin levels and mortality were compared.Results For patients with high level obstruction,incidence of fever in ENBD+ERBD group and double ERBD group were significantly higher than that of ENBD group (1/18 VS 4/10,P ＜ 0.05;0 VS 4/10,P ＜ 0.05).There were no significant differences in incidence of fever(1/18 VS 0) or bilirubin level decrease (17/18 VS 14/15) between ENBD+ERBD group and double ERBD group.For high level obstruction,if the patient had biliary imaging with contrast medium during operation,they would have higher incidence of high fever[(4/10 VS 3.45％ (1/28)] and mortality(3/11 VS 0),lower declining rate of bilirubin level [10/14 VS 96.55％(28/29)] than those without.Conclusion Both left and right hepatic ducts drainage should be recommended for high level biliary obstruction.During the operation,contrast medium should be limited to minimum dose,and should be avoided in high level obstruction cases.Air biliary imaging could be used when necessary.

Objective The present study investigated the expression of heat shock cognate protein 70 (Hsc70) in the synovial tissues and blood samples of patients with rheumatoid arthritis (RA) to determine the pathological role of this protein in the pathogenesis of the disease.Methods The expression of Hsc70 in synovial membranes was quantitatively analyzed by immunohistochemistry,real-time quantitative PCR and western blotting.The samples from osteoarthritis (OA) and ankylosing spondylitis (AS) were used as controls.The levels of Hsc70 in blood of patients with RA were determined using enzyme linked immunosorbent assay (ELISA) with the samples of the healthy subjects as controls.Statistical analysis was conducted with one-way ANOVA,LSD test and Spearmen's correlation.Results Immunohistochemistry showed that Hsc70 had significantly increased expression in synovial tissues of RA than in the samples of OA and AS.Real-time PCR and western blotting confirmed the above findings.ELISA detected significantly elevated level of Hsc70 in blood of patients with RA as compared with samples from the controls (P＜0.01).Conclusion The study suggests that the up-regulation of Hsc70 may be involved in the pathogenic process of RA.

Objective To evaluate angiogenesis of the benign and malignant solid thyroid nodules with color Doppler ultrasound and immunohistochemistry staining. Methods Fifty-six solid thyroid nodules in 55 patients (28 papillary thyroid cancer, 23 goiter, 4 adenoma, 1 Hashimoto' s disease) were observed before surgery with color Doppler ultrasound. Pathological specimens of paraffin-embedded were immunohistochemically stained with CD34 and VEGF antibody. Results There were significant differences between the benign and malignant thyroid nodules in vascular morphology and regional rich blood flow. The irregular or less irregular vessels were found in 75 % of the malignant nodules. Regional rich blood flow or suspicious regional rich blood flow were found in 64. 3% of malignant nodules. The regular vessels were found in 89. 3% of the benign nodules, non-regional rich blood flow was found in 71.4% of the benign nodules. The number of CD34 in malignant lesions [(37.31 ± 11.55)/HP] was significantly higher than benign lesions [(29. 02 ± 8.32)/HP, P = 0.04]. There was a significantly difference of VEGF expression between the benign and malignant nodules which was higher in malignant nodules than in benign nodules(P ＜ 0.01). Conclusions Compared with the benign nodules, the vessles in malignant thyroid nodules were irregular,the distribution of vessles was asymmetry and angiogenesis was active.

Objective To evaluate the contrast enhanced perfusion pattern of PTC micro-vascular imaging (MVI) quantitatively.Investigate the correlation between PTC MVI features and CD34 micro-vascular density (MVD).Methods Thirty-nine pathological and clinical confirmed sporadic PTCs were evaluated with real-time gray-scale contrast-ernhanced micro-vascular imaging under a low mechanical index.The micro-bubble agent was SoneVue.Of the 39 PTCs,33 were classical PTCs,6 were PTC with follicular variant (FVPTC).The △ ROI,which is the subtraction of peak echo intensity between the lesion region of interest (ROI) and normal thyroid parenchyma ROI,was used to evaluate the perfusion characteristics of PTC MVI quantitatively.The paraffined specimens were selected for immunohistochemical (IHC) staining for CD3,and the correlation between △ ROI and the CD34 were evaluated.Results △ ROI was strongly correlated with the CD34 expression (P=0.000),significant differences were detected in the distribution pattern of △ ROI value among different CD34 expression levels,no overlapping of the mean △ ROI values and the 95％ confidence intervals was found among the 3 CD34 expression levels.The PTC MVI perfusion was classified into 3 patterns,low perfusion,focal perfiusion and high perfusion,on the basis of combining△ ROI values with the peak ehco pattem in time-intensity curve.Conclusions The △ ROI is strongly correlated with the CD34 expression in papillary thyroid cancer.It can be used for the quantitative evaluation ofPTC MVI pattem and intensity as an objective indicator.

Pulmonary vein isolation (PVI) is a new ,effective and radical method to cure atrial fibrillation .Within a period after radiofrequency ablation (RFA) ,coagulation system is activated in patients ,then thrombus incident such as cerebral embolism may happen .The present article made a review on its mechanism .

Objective To investigate the efficacy of endoscopy for the treatment of benign biliary stricture after biliary surgery.Methods The clinical data of 127 patients with benign biliary stricture after biliary surgery at the First Hospital of Lanzhou University from January 2007 to December 2011 were retrospectively analyzed.According to the Bismuth classification,there were 60 patients with type Ⅰ,35 with type Ⅱ,21 with type Ⅲ and 11 with type Ⅳ.The efficacies of endoscopy for the treatment of biliary stricture with different Bismuth subtypes were analyzed.Results The location and severity of biliary stricture were confirmed by endoscopic retrograde cholangiopancreatography (ERCP) + cholangiography.Sixteen patients ( including 7 with type Ⅲ and 9 with type Ⅳ) were transferred to surgical treatment due to severe biliary stricture.A total of 111 patients underwent endoscopic treatment successfully,with the success rate of 87.4％ (111/127).The success rates of endoscopy for the treatment of patients with Bismuth Ⅰ,Ⅱ,Ⅲ and Ⅳ biliary strictures were 95％ (57/60),86％ (30/35),9/14and 1/2,respectively.Twenty-nine patients were implanted with retrievable metallic biliary stent,and 82 were implantated with plastic biliary stent.Of the 111 patients,only 6 patients were complicated by acute pancreatitis,and they were cured by conservative treatment.The alleviative rates of yellow skin and icteric sclera,tenderness and distending pain of right upper quadrant,fever were 73％ (81/111 ),83％ (74/89),90％ (73/81 ) and 89％(68/76) at 1 week after treatment,and they were 88％ (98/111),91％ (81/89),94％ (76/81) and 92％(70/76) at 8 weeks after treatment.The efficacy of endoscopy was good in 97 patients and poor in 14 patients,and the 14 patients were converted to open surgery.The symptoms including yellow skin and icteric sclera,tenderness and distending pain of right upper quadrant,fever were completely alleviated at postoperative month 6.Conclusion Endoscopic treatment for benign biliary stricture is safe and effective.

Background and purpose:Three-dimensional power Doppler angiography (3D-PDA) is a new technique to investigate the vessels in the organs, but the research in thyroid is limited. The purpose of this research was to investigate three-dimensional power Doppler angiography (3D-PDA) in differentiating malignant from benign thyroid nod-ules.Methods:This study prospectively evaluated 103 lesions in 94 patients who were scheduled for surgery. The patients underwent preoperative 3D-PDA scanning. Analysis of the 3D-PDA characteristics includes blood flow pattern, the num-ber of blood vessels, the shape of vessels, the spatial distribution of the vessels, the existence of rich local blood flow within nodules or in the parenchyma surrounding the nodules. This study also analyzed the difference between the benign lesions and the malignant lesions.Results:There were 50 benign lesions and 53 malignant lesions. The sensitivity and specificity of irregular vessels, the asymmetry spatial distribution, rich local blood flow within nodules or in the parenchyma surround-ing the nodules were 64.2%, 96.0%; 56.0%, 88.0%; 54.7%, 96.0%; 60.4% and 94.0%, respectively. The sensitivity, speci-ficity, positive predictive value, negative predictive value and accuracy of 3D-PDA were 83.0%, 94.0%, 93.6%, 83.9% and 90.3%, respectively.Conclusion:3D-PDA provides a useful tool to investigate vascularization of thyroid leisions.This technique is feasible for clinical application and plays an important role in diagnosis of thyroid nodules.

Objective To investigate the efficacy of combined application of laparoscope and multiple endoscopes for the treatment of extra-hepatic bile duct stones.Methods The clinical data of 3780 patients with extra-hepatic bile duct stones who were admitted to the First Hospital of Lanzhou University from March 1998 to June 2010 were retrospectively analyzed.According to the condition of bile duct stones,laparoscope,choledochoscope and duodenoscope were applied separately or combinately.All patients were divided into A,B and C groups.Patients in group A were treated by laparoseopy,choledochoscopy or duodenoscopy;patients in group B were treated by choledochoscopy+duodenoscopy.duodenoscopy+laparoscopy or laparoscopy+choledochoscopy;patients in group C were treated by laparoscopy+duodenoscopy+choledochoscopy.The efficacies of different treatment approaches were analyzed by comparing the results of imaging examination and follow-up.Results The curative rate and complication rate of the group A were 89.54%(1276/1425)and 6.73%(86/1276),respectively.Of the 149 patients in the group A who were failed in the treatment,83 patients were transferred to the group B,and 66 patients were transferred to the group C.The curative rate and complication rate of the group B(including 83 patients transferred from the group A)were 95.93%(1719/1792)and 4.07%(70/1719),respectively,and 73 patients who were failed in the treatment were transferred to the group C.The curative rate and complication rate of the group C(including 139 patients transferred from the group C)were 99.75%(783/785)and 0.26% (2/783),respectively,and 2 patients who were failed in the treatment received open surgery.Conclusion Combined application of laparoscope and endoscopes could raise the success rate of stone clearance and decrease the postoperative complications.

Objective To summarize our experience of laparoscopic surgery for complex choledochal cysts (type Ⅳ-A). Methods The clinical data of 65 children of choledochal cyst undergoing laparoscopic choledochal cyst resection were retrospectively reviewed from 2002 to 2009 in our institute.Among those type Ⅳ-A cyst was found in 16 patients. Hepaticojejunostomy was performed using a Roux-en-Y jejunal loop after extrahepatic cyst excision and ductoplasty. Results Laparoscopic procedures were successfully performed in 16 patients with type Ⅳ-A cysts. The stenotic segment was splited or excised and a wide hepaticojejunostomy was completed at the porta hepatis in 8 patients with a stricture extending to the level of common hepatic duct. The constrictive confluence of the bilateral hepatic duct was incised and the bi-ductal cystojejunostomy was achieved at the bifurcation in 4 cases. A septum was found at the orifice of right hepatic duct and was excised through the hilar stoma in 2 cases. A downstream stricture of the left hepatic duct was incised from the hilum to the dilated segment along the lateral wall in 2 patients, so that a long intrahepatic cystojejunostomy was completed in an oblique course. Postoperative complications developed in 2 cases including temporary bile leakage in one case and anastomotic stricture in another. The intrahepatic cysts were remarkably reduced in size during the follow-up. Conclusions With the magnified laparoscopic view, the radical resection of extrahepatic cyst and correction of the intrahepatic bile ductal stenosis can be easily performed. Laparoscopic hepaticojejunostomy and/or intrahepatic cystojejunostomy is effective and safe for children with type Ⅳ-A choledochal cysts.