Streptokinase is an extracellular protein, extracted from certain strains of beta hemolytic streptococcus. It is a non-protease plasminogen activator that activates plasminogen to plasmin, the enzyme that degrades fibrin cloth through its specific lysine binding site; it is used therefore as a drug in thrombolytic therapy. The rate of bacterial growth and streptokinase production was studied in condition of excess glucose addition to culture media and its pH maintenance. The streptokinase product of the bacterial culture was preliminary extracted by salt precipitation and then purified by affinity chromatography on plasminogen substituted sepharose-4B in a condition that the plasminogen active site was protected from streptokinaseinduced activation. The purity of streptokinase was confirmed by SDS-PAGE and its biological activity determined in a specific streptokinase assay. The results showed that in the fed –batch culture, the rate of streptokinase production increased over two times as compared with the batch culture while at the same time, shortening the streptokinase purification to a single step increased the yield over 95 % at the chromatography stage.

Introduction: Lipoprotein (a) [Lp (a)] is one of the cholesterol-rich plasma lipoproteins, which is a low-density lipoprotein-like particle. Lp (a) displays strong athero-thrombotic properties. In Lp (a), apoprotein (a) [Apo (a)] is covalently linked by a single disulfide bridge to apoproteinB100 (apoB100). Apo (a) is a glycosylated protein similar to plasminogen. Plasminogen is a fibrinolytic proenzyme with kringle-like sequences. Lipoprotein (a) in human plasma appears as four subspecies based on its affinity for lysine sepharose: Lp (a) Lys-, Lp (a) Lys+1, Lp (a) Lys+2 and Lp (a) Lys+3. Pathogenesis of Lp (a) as a risk factor for cardiovascular disease may depend on its lysine-binding site activity. It is suggested that Lp (a) LYS+1, Lp (a) Lys+2 and Lp (a) Lys+3 have, athero-thrombotic properties. In Iran there have not been many studiesori Lp (a) subspecies. Therefore the aim of this study was to separate Lp (a) subspecies according to their affinity for lysine, using lysine sepharose affinity chromatography. Methods: We separated four different subspecies of Lp (a) by changing pH and molarity of the mobile phase. 100 ml of pooled serum was collected.Results: The total Lp (a) was 46 mg and composed of 12 mg (24%) Lp (a) Lys-, 4 mg (8%), Lp (a) LYS+1, 23 mg (50%), Lp (a) Lys+2, and 4 mg (8%) Lp (a) Lys+3.Electrophoretic mobility of four different subspecies of Lp (a) was assessed on polyacrylamide disk electrophoresis. Purity of the separated fraction was confirmed and there was no difference in electrophoretic mobility between the four subspecies of Lp (a).Discussion: The ability to separate Lp (a) subspecies will enable us to investigate factors affecting atherosclerosis and cardiovascular diseases by studying their effect on Lp (a) subspecies in the future.

Two forms of Acetylcholinesterase (AChE) could be found in the Wistar rat brain homogenate, soluble and membrane bound enzyme. In order to study and purification of this enzyme, a homogenate of Wistar rat brain was prepared in a phosphate buffer, after centrifugation, supernatant which contains the soluble enzyme was separated. In the second stage the pellet was resuspended in the same volume of phosphate buffer contained Triton X-100 and centrifuged again. The supernatant this time contained the membrane bound enzyme. This enzyme was isotaled and purified using MAP-Agarose.

Alpha toxin, sometimes referred to as phospholipase C (PLC), is a significant toxin generated by C. perfringens that possesses both deadly and dermonecrotic properties. All strains (A, B, C, D, E) of C. perfringens generate varying quantities of this toxin, which is recognized as a key various  factor in the development of clostridial myonecrosis.Alpha-toxin is known to significantly influence several human and animal disorders, exhibiting deleterious impacts, specifically on the intestinal system. The primary objective of this research was to conduct a comparative analysis of two distinct purification techniques in order to extract alpha-toxin from C. perfringens type A. The first approach involved the alpha-toxin purification through a series of techniques, including ammonium sulfate precipitation, ion-exchange chromatography using DEAE Sephadex at pH values of 7 and 9, and gel filtration chromatography using Sephadex G-100, while the second approach was implemented using the affinity chromatography. The degree of purity of alpha-toxin was assessed at each stage of the purification process by the utilization of SDS-PAGE.The protein content and hemolysis activity were also quantified at each purification step. Based on the obtained findings, examining these two approaches revealed that the first method yielded a percentage of 88, while the second method yielded percentage of 91.7. The specific activity values obtained from the calculations for the first and second methods were 69170 U/mg and 105.71 U/mg, respectively. Our research shows that affinity chromatography can produce a highly pure alpha toxin with a specific activity of 108700 (HU/mg), produced by Clostridium perfringens type A.

Background and Objectives Hemophilia B is a genetic disorder due to deficiency or complete absence of factor IX coagulation factor. Treatment of choice for these patients is use of factor IX concentrates. Therefore, purification of plasma proteins and separation of factor IX have been major objectives for scientists involved in this field. In this respect, purification procedure using ion exchange chromatography is widely used, but in the past decade affinity chromatography was also introduced. The objective of the present study has been to apply both techniques for the purification of factor IX and compare the quality and yield of the product. Materials and Methods For the purification procedure, chromatography columns (XK-16), containing DEAE sepharose and Heparin sepharose were used. Factor IX coagulation activity was measured using a one-stage coagulation assay and factor IX antigen was quantified using ELISA technique.  Results The specific activity and relative increase in purity of factor IX was calculated and it was demonstrated that specific activity improved from 3.1 IU/mg using DEAE ion exchange to 29 IU/mg when affinity chromatography was added and purity was increased from 155 to 1450 respectively. Conclusions The present study demonstrates that addition of an affinity chromatography step using heparin sepharose is a major improvement in the purification of factor IX, where both specific activity and purity are increased considerably.  

Vero toxins are a group of closely related subunit toxins, that have significant effects in pathogenesis of verotoxigenic Escherichia coli. Recent evidences suggest that vero toxinl induce cytotoxic effects on some tumor cell lines. So isolation of verotoxigenic Escherichia coli strains and toxin purification are very important subjects. Inspite of different genetically desigened methods in detection of vera toxigenic Escherichia coli strains ,serological detection with polyclonal anti-verotoxinantibodies is a very rapid and important method.Ourpurpose ispurification ofverotoxinl from screened diarrheagenic strains, to preparation of serologicalldt in the future.Toxin production in five diarrehagenicstrains was screened by Vero toxin-producing Escherichia coli-Reverse passive latex agglutination test (VTEC-RPLA kit, Oxoid, Co, Ltd) and then purified by affinity chromathgraphy of receptor glycolipid (PI blood group antigen).Screened strain produced large amounts of toxin that have been comparable with standard toxin. Electrophoresis of toxin indicate an intense and sharp band in rage of 70 kilodalton. Intraperitoneal injection in Balb/c have several effects such as hind-legparalysis, cachexiasand diarrheae.In addition in exposed mouse and Hela and Vera cell lines teratment,we observed severe cytotoxic effects.

Introduction: Egg yolk is a rich and available source of immunoglobulin Y (IgY) that can be used in medical diagnosis and treatment against microbial agents.Methods and Materials: In this study anti-human IgG specific IgY was produced in hens and purified from egg yolk using three different methods. After immunization of hens against human IgG, IgY was extracted from egg yolk by the acidic water dilution method and purified using three methods; polyethylene glycol precipitation, ion exchange chromatography and affinity chromatography. Molecular mass and activity of the purified antibody were estimated using SDS-PAGE and ELISA, respectively.Results: The results showed that the affinity produced IgY antibodies has purity and yield of 95% and 78% respectively. In addition, the suitable and simple method for purification of IgY fraction was prepared by PEG precipitation with recovery and purity of 90% and 95% respectively. Ion exchange chromatography product had recovery and purity of 72% and 68%. Furthermore the molecular weight (MW) of intact IgY and its light and heavy chains were estimated to be 190, 27 and 67 KD, respectively. This product can be used for diagnosis of IgG in different diagnostic tests.