RESUMEN
【Objective】 To establish a new method for the determination of fibrinogen content in cryoprecipitated antihemophilic factor. 【Methods】 Fibrinogen (Fib) could bind with sheep anti-human fibrinogen (anti-Fib) specifically and further form antigen-antibody complex. When the Fib was present in the solution, the fluorescence of fluorescein isothiocyanate (FITC) labeled on the anti-Fib (FITC-anti-Fib) was quenched due to the formation of immune complex. The fluorescence quenching degree of FITC-anti-Fib was positively correlated with Fib concentration (cFib) in a certain concentration range. 【Results】 The linear relationship between fluorescence quenching degree [(I0-I)/I0] of FITC-anti-Fib and ln(cFib) was (I0-I)/I0=15.53ln(cFib)+ 80.79 (R2=0.99) when the cFib was in the range of (0.007 8-0.560 0) g/L. The recovery of Fib was (96.77-102.43) %. When the method was applied to determine Fib at high, medium, and low concentrations, the obtained intra-day variation coefficients were 0.31%, 0.56%, and 0.49%, respectively, and the inter-day variation coefficients were 3.81%, 3.06%, and 4.13%, respectively. There was no significant difference between the results measured by fluorescence quenching method and coagulation method (t=-0.075, P>0.05). 【Conclusion】 In this work, a new fluorescence method for the determination of Fib in cryoprecipitated antihemophilic factor was successfully established based on the specific combination of fib and FITC-anti-Fib. The method is simple and rapid. The obtained results were accurate and reliable by using this method to determine Fib.
RESUMEN
In bleeding events caused by various reasons, coagulation dysfunction due to massive consumption or loss of coagulation factors is often life-threatening. In recent years, medical evidence of using human fibrinogen concentrate to prevent or treat bleeding has emerged, providing more options for clinical treatment of bleeding. In this paper, the stability, pharmacokinetics and safety of human fibrinogen, as well as the latest advances in its clinical application in obstetrics, cardiac surgery and traumatic massive hemorrhage were summarized, and new reasonable suggestions for clinical application of human fibrinogen were also put forward.
RESUMEN
Background: Studies evaluating the hemostatic effects of fibrinogen administration in cardiac surgery are not conclusive. Aims: We investigated whether the use of a low-dose human fibrinogen in case of clinical bleeding after protamine administration and concomitant low FIBTEM values is effective in reducing postoperative bleeding. Secondary end-point was to investigate the consumption of allogeneic blood products. Setting and Design: This was a retrospective matched study conducted at university hospital. Materials and Methods: Among 2257 patients undergoing surgery with cardiopulmonary (CPB) bypass, 73 patients received a median dose of 1 g human fibrinogen (ROTEM-Fibri group). This group was matched with 73 patients who had not received human fibrinogen (control group) among 390 patients having undergone surgery at the moment FIBTEM analysis was unavailable. Statistical Analysis: Matching was performed for the type and the presence of redo surgery. McNemar and Wilcoxon paired tests were used to respectively compare the categorical and quantitative variables. Results: The CPB bypass time was significantly higher in the ROTEM-Fibri group (P = 0.006). This group showed significantly higher bleeding in the first 12 and 24 h postoperatively (P < 0.001) and required significantly more transfusion of blood products (P < 0.001) and surgical revision (P = 0.007) when compared with the control group. There was no significant difference in the number of thromboembolic complications. Conclusions: These results show that the administration of 1 g of fibrinogen based on low-FIBTEM values and clinical bleeding after protamine administration does not stop bleeding and the need for transfusion of allogeneic blood products.
RESUMEN
Objective To construct a eukaryotic expression vector in Pichia pastoris containing human fibrinogen gene, in order to achieve high level secretory expression in extracellular.Methods Expression plasmid,pGAPZαA-FGB-FGG-FGA-AOX1,was constructed by inserting the synthesized sequence encoding human fibrinogen(FGA, FGB,FGG) and then introduced into Pichia pastoris SMD1168H by electroporation.Transformants were availably screened by Zeocin resistance,the expression of recombinant protein was identified by SDS-PAGE and Western blot analysis, the protein yield was tested by ELISA assay.After ultrafiltration and purification, the biological activity of protein was detected.Results The crude yield of human fibrinogen in Pichia pastoris supernatant reached 15 mg/L in flask and the biological aggregation activity was determined.Conclusion The human fibrinogen gene was obtained and successfully expressed in Pichia pastoris and the active products were secreted into the medium.
RESUMEN
OBJECTIVE: To establish a quantitative determination method of sucrose in human fibrinogen by HPLC. METHODS: An HPLC method was developed to specifically determine sucrose on Zorbax carbohydrate analysis column with Waters Alliance system and 2414refractive index detector. The separation was performed at 30°C using a mobile phase consisting of acetonitrile and water (70:30) at a flow rate of 1.4 mL · min-1. Thirteen batches of recombinant human coagulation factor VM samples were selected for methodology comparison of the established HPLC method with the IEC-HPLC method adopted by Ch. P 2010, because these samples only contained sucrose and did not receive dry-heat treatment. RESULTS: The RSDs (n=6) of the retention time and peak area were 0.17% and 0.09%, respectively. The recoveries of sucrose at low (5 mg · mL-1), middle (10 mg · mL-1), high (15 mg · mL-1) concentration were 96.2%, 98.8% and 100.3%, respectively. The average recovery was 98.4% and the linear correlation coefficient r was 1.0000 in the ranges of 2-20 mg · mL-1. Statistical analysis showed that the amounts of sucrose in 13 batches of recombinant human coagulation factor VIII samples determined by the HPLC method and IEC-HPLC method had no significant difference (P≤0.05) and the correlation coefficient r was 1.0000. CONCLUSION: The proposed HPLC method is simple, accurate and re-peatable for determination of sucrose in dry-heat treated human fibrinogen product. The HPLC method showed good accordance with IEC-HPLC method for assay results of sucrose, thus can be widely applied.
RESUMEN
PURPOSE: To find the most effective treatments in cystic diseases of soft tissue using three different treatment modalities. MATERIALS AND METHODS: We performed three different treatment modalities-curative excision, incision & steroid injection, and aspiration & human fibrinogen injection-in 75 patients from June 1997 to May 1998 and followed up for at least one year. We analyzed the recurrence rate and patients' satisfaction according to their treatment modalities. We used the Fischer's exact test (P=0.05) to evaluate the results. RESULTS: In curative excision 4 of 23 patients (18%) were recurred, 6 of 14 patients (43%) were recurred in incision & steroid injection, and 20 of 36 patients (56%) in aspiration & human fibrinogen injection, and it was meaningfully different in statistics. 16 of 23 patients (70%) were satisfied with the result in curative excision, 4 of 14 patients (29%) in steroid injection, and 11 of 36 patients (31%) in aspiration & human fibrinogen injection, and it was also different in statistics. Recurrence was the most popular reason of dissatisfaction. CONCLUSION: This study showed that curative excision was the most effective cure method. Because both incision & steroid injection and aspiration & human fibrinogen injection method were easier to perform, cosmetically preferable and much cheaper in cost, these two methods would be recommended before surgical intervention.