Evaluation of Human Anti Igg Polyclonal Antibody Production Conjugated with Peroxidase in Egg Yolk

2020 ◽  
Author(s):  
Fahimeh ABDOLMALEKI ◽  
Zahra ZAMANI ◽  
Somayeh TALEBI
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Polyclonal Antibody ◽  
Antibody Production ◽  
Polyclonal Antibodies ◽  
Egg Yolk ◽  
Elisa Test ◽  
Heavy Chains ◽  
University Of Technology ◽  
Sds Page

Background: Egg yolk is a rich and accessible source of yolk immunoglobulin (Y immunoglobulin). Presently, polyclonal antibodies from mammalian sources are used for diagnosis. Antibody production from egg yolk gives a higher yield and turnover than that from lab animals, and invasive methods such as phlebotomy and causing stress to the animals are not required. Due to the issues regarding mammalian antibodies, we aimed to evaluate the human anti-IgG polyclonal antibody production conjugated with peroxidase in egg yolk.Methods: Population of laying hens reared in Agriculture/Isfahan University of Technology were used in 2017. After immunizing hen against pure human IgG, specific IgY (yolk immunoglobulin) was purified from the yolk by sedimentation with polyethylene glycol (PEG6000). To assess the molecular weight and activity of the product, SDS-PAGE and ELISA-test were used, respectively.Results: The complete molecular weight of IgY was 180 kDa and the molecular weight of its light and heavy chains were 27 and 67 kDa, respectively.Conclusion: Antihuman IgG IgY had a purity above 90%. The product of this study can be used to measure IgG class antibodies in order to diagnose different diseases.

Catabolism of streptokinase and polyethylene glycol-streptokinase: evidence for transport of intact forms through the biliary system in the mouse

Blood ◽  
1990 ◽  
Vol 76 (1) ◽  
pp. 73-79 ◽  
Author(s):  
FH Brucato ◽  
SV Pizzo
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Gel Electrophoresis ◽  
Proteinase Inhibitors ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gi Tract ◽  
Substantial Fraction ◽  
Sds Page ◽  
Derivatives Of

Abstract The catabolism of streptokinase (SK) and polyethylene glycol derivatives of SK (PEG-SK) were studied in mice. The clearance and catabolism of SK:plasmin (SK:Pm) and PEG-SK:Pm activator complexes were also investigated. Native 125I-SK cleared rapidly (t1/2 = 15 minutes) from the circulation, with the majority of the ligand accumulating in the liver and gastrointestinal (GI) tract and a substantial fraction also localizing in the kidneys. SK, which was removed from the plasma by the liver, was secreted into bile and then the GI tract. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that 125I-SK recovered from liver and bile was homogeneous and of the same molecular weight (mol wt approximately 50,200) as native SK. PEG-125I-SK cleared slowly (t1/2 greater than 200 minutes), with more than 80% of the preparation localizing in liver and GI tract. The PEG-125I-SK secreted into the bile was also intact. The bile containing 125I-SK was incubated with stoichiometric amounts of plasminogen and electrophoresed under nondenaturing conditions. This study demonstrated that the secreted SK was able to form SK:Pg complexes. SDS-PAGE also showed activation of 125I-Pg that was incubated with recovered bile containing the SK. 125I-SK:Pm catabolism was also studied. In these experiments, the mol wt approximately 42,000 fragment obtained when SK is cleaved by plasmin was found in the bile. This fragment of 125I-SK was not recovered as part of a complex with plasmin, consistent with our previous observations that catabolism of SK:Pm involves transfer of the plasmin to plasma proteinase inhibitors while SK is catabolized independently. By contrast, when PEG-125I-SK:Pm was injected into mice, only intact PEG-125I-SK was found in the bile, consistent with our previous observations that the PEG derivatization blocks its degradation by plasmin.

scholarly journals The Potential of Candida Biofilm Protein as Bioreceptor for Candidiasis Immunoassay

2018 ◽  
Vol 14 (1) ◽  
pp. 47 ◽  
Author(s):  
Masfufatun Masfufatun ◽  
Loo Haryanto ◽  
Harsono Harsono
Keyword(s):  
Molecular Weight ◽  
Candida Albicans ◽  
Polyclonal Antibody ◽  
Control Group ◽  
Potential Candidate ◽  
Dot Blot ◽  
Protein Extract ◽  
Treatment Groups ◽  
Sds Page ◽  
Mechanical Methods

Abstract: Candidiasis or infection that is caused by Candida has become a new list of the therapeutical problems recently. The difficulties in diagnosing are the main cause of the unsatisfactory results from common therapies and diagnosis methods. This has urged researchers to find alternative ways in candidiasis diagnosis such as serology-based detection using antigen or antibody development. The aim of this study was to evaluate the potential of protein derived from Candida albicans biofilm as bioreceptor on candidiasis immunoassay through Dot Blot method. The research method used descriptive method with the following stages: (1) preparation of Candida albicans biofilm (2) extraction of Candida albicans protein through enzymatic and mechanical methods, (3) determination of protein molecular weight with SDS-PAGE (4) production of polyclonal anti- candida and (5) analysis of protein extract as bioreeceptor on dot blot. Profile of biofilm proteins on SDS-PAGE analysis were shown on molecular weight 27,42; 29,89; 38,10; 44,90; 48,75; 52,92; 55,14; 59,86; 70,56; 87,36; 102,54;115,05; 130,14;143,14;181,53 kD. There were differences in the intensity of dots in the control group (44070) and treatment groups (63170.5). It is noticeable that biofilm protein extract of C. albicans can be used for induction of anti-Candida polyclonal antibody production as the potential candidate of bioreceptor in candidiasis immunoassay. Keywords: SDS-PAGE, polyclonal antibody, immunoassay, dot blot, biofilm

scholarly journals Immunodetection of Furcraea Necrotic Streak Virus-FSNV in Fique Plants (Furcraea Macrophylla Baker) Using A Polyclonal Antibody IgY Produced in Chicken Egg Yolk

2021 ◽  
Author(s):  
Deisy Toloza-Moreno ◽  
Laura Villamizar-Rivero ◽  
Paola Cuartas-Otálora ◽  
Gloria Barrera-Cubillos
Keyword(s):  
Early Detection ◽  
Polyclonal Antibody ◽  
Polyclonal Antibodies ◽  
Egg Yolk ◽  
Primary Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antibody Concentration ◽  
Streak Virus ◽  
Dot Blot ◽  
Specificity And Sensitivity

Abstract The necrotic streak of the fique (Furcraea spp.) or “Macana” disease is considered the most limiting disease for this crop in Colombia, whose causal agent is the Furcraea Necrotic Streak Virus - FSNV (RNA +). Currently, there are no strategies to control the disease, being necessary to develop methods for early detection of this disease in the planting material before being taken to the field. In this study, polyclonal antibodies produced in egg yolk (IgY) were assesses for detection FSNV. Two immunoenzymatic methodologies were standardized: Dot Blot Immunobinding Assay (DBIA) and Enzyme Linked Immunosorbent Assay (ELISA), determining their specificity and sensitivity. The detection limit by DBIA corresponded to 8 µg/mL of purified virus suspension using 10 µg/mL of primary antibody. In the ELISA test, the primary antibody concentration of 3 µg/mL (1:800 dilution) detected the antigen at concentrations between 10 and 70 µg/mL. The polyclonal antibody anti-FNSV IgY allowed the detection of FNSV in samples of purified virus and extracts of roots and leaves of fique plants with symptoms of “Macana” disease and did not produce any signal with the control samples. Results showed the potential of using egg yolk IgY for immunological test as a novel approach for the early detection of FNSV in fique plants.

scholarly journals Catabolism of streptokinase and polyethylene glycol-streptokinase: evidence for transport of intact forms through the biliary system in the mouse

Blood ◽  
1990 ◽  
Vol 76 (1) ◽  
pp. 73-79
Author(s):  
FH Brucato ◽  
SV Pizzo
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Gel Electrophoresis ◽  
Proteinase Inhibitors ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gi Tract ◽  
Substantial Fraction ◽  
Sds Page ◽  
Derivatives Of

The catabolism of streptokinase (SK) and polyethylene glycol derivatives of SK (PEG-SK) were studied in mice. The clearance and catabolism of SK:plasmin (SK:Pm) and PEG-SK:Pm activator complexes were also investigated. Native 125I-SK cleared rapidly (t1/2 = 15 minutes) from the circulation, with the majority of the ligand accumulating in the liver and gastrointestinal (GI) tract and a substantial fraction also localizing in the kidneys. SK, which was removed from the plasma by the liver, was secreted into bile and then the GI tract. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that 125I-SK recovered from liver and bile was homogeneous and of the same molecular weight (mol wt approximately 50,200) as native SK. PEG-125I-SK cleared slowly (t1/2 greater than 200 minutes), with more than 80% of the preparation localizing in liver and GI tract. The PEG-125I-SK secreted into the bile was also intact. The bile containing 125I-SK was incubated with stoichiometric amounts of plasminogen and electrophoresed under nondenaturing conditions. This study demonstrated that the secreted SK was able to form SK:Pg complexes. SDS-PAGE also showed activation of 125I-Pg that was incubated with recovered bile containing the SK. 125I-SK:Pm catabolism was also studied. In these experiments, the mol wt approximately 42,000 fragment obtained when SK is cleaved by plasmin was found in the bile. This fragment of 125I-SK was not recovered as part of a complex with plasmin, consistent with our previous observations that catabolism of SK:Pm involves transfer of the plasmin to plasma proteinase inhibitors while SK is catabolized independently. By contrast, when PEG-125I-SK:Pm was injected into mice, only intact PEG-125I-SK was found in the bile, consistent with our previous observations that the PEG derivatization blocks its degradation by plasmin.

scholarly journals PERAN PROTEIN PILI 11 kDa Streptococcus pneumoniae SEBAGAI PROTEIN HEMAGLUTININ DAN ADHESIN

2020 ◽  
Vol 7 (1) ◽  
Author(s):  
Diana Chusna Mufida ◽  
Yuna Annisa Salsabila ◽  
Enny Suswati ◽  
Bagus Hermansyah ◽  
Dini Agustina
Keyword(s):  
Streptococcus Pneumoniae ◽  
Epithelial Cells ◽  
Immune Responses ◽  
Western Blotting ◽  
Polyclonal Antibody ◽  
Polyclonal Antibodies ◽  
Pearson Correlation ◽  
P Value ◽  
Sds Page ◽  
Pearson Correlation Test

Role of Pili Protein 11 kDa of Streptococcus pneumoniae as Hemagglutinin and Adhesin Protein Streptococcus pneumoniae has pili which play roles in adhesion, colonization of nasopharyngeal epithelial cells, and phagocytic inhibition of immune cells. This study aimed to determine the characteristics of the 11 kDa pili protein as hemagglutinin and adhesin, as well as their immune responses. The 11 kDa pili protein from S. pneumoniae was isolated by SDS-PAGE, purified by electroelution and dialysis. Hemagglutination and adhesion tests were carried out on the protein, and western blotting of the polyclonal antibody immune responses were evaluated. Hemagglutination test showed that the 11 kDa pili protein played a role in the hemagglutination process up to 2-time dilution. Adhesion test showed there was a correlation between the dose of the protein and the bacteria attached to the epithelial cells. The Pearson correlation test showed a P value of 0.010 and a correlation coefficient of R = -90.919. Quadratic regression test produced R2 = 0.974. Western blotting test showed that 11 kDa pili protein polyclonal antibodies recognized 67 kDa and 11 kDa pili proteins. The study concluded that the 11 kDa S. pneumoniae pili protein acted as hemagglutinin and adhesin, and the polyclonal antibody protein responded to 67 pDa and 11 kDa BM pili proteins.Keywords: adhesin, hemagglutinin, pili, protein 11 kDa, Streptococcus pneumoniae ABSTRAKStreptococcus pneumoniae memiliki pili yang berperan dalam adhesi, kolonisasi sel epitel nasofaring, serta sebagai inhibitor fagositosis sel imun. Penelitian ini bertujuan mengetahui karakteristik protein pili 11 kDa sebagai hemagglutinin dan adhesin serta respons imunnya. Protein pili 11 kDa dari bakteri S. pneumoniae diisolasi secara SDS-PAGE, dipurifikasi dengan elektroelusi dan dialysis. Uji hemaglutinasi dan adhesi dilakukan pada protein tersebut, serta dievaluasi respon imun poliklonal antibodinya secara western blotting. Uji hemaglutinasi menunjukkan protein pili 11 kDa berperan dalam proses hemaglutinasi hingga pengenceran 2 kali. Uji adhesi menunjukkan korelasi antara dosis protein dan bakteri yang menempel pada sel epitel. Uji korelasi Pearson menunjukkan P value 0,010 dan koefisien korelasi R = -0,919. Uji regresi Quadratic menghasilkan R2 = 0,974. Uji Western blotting menunjukkan antibodi poliklonal protein pili 11 kDa mengenali protein pili 67 kDa dan 11 kDa. Penelitian ini berkesimpulan protein pili 11 kDa S. pneumoniae berperan sebagai hemaglutinin dan adhesin, serta antibodi poliklonal protein tersebut memberi respons terhadap protein pili BM 67 kDa dan 11 kDa. 

scholarly journals Chemical Modification of Sweet Potato β-amylase by Mal-mPEG to Improve Its Enzymatic Characteristics

Molecules ◽  
2018 ◽  
Vol 23 (11) ◽  
pp. 2754
Author(s):  
Xinhong Liang ◽  
Wanli Zhang ◽  
Junjian Ran ◽  
Junliang Sun ◽  
Lingxia Jiao ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Sweet Potato ◽  
Specific Activity ◽  
Potato Starch ◽  
Sds Page ◽  
Significant Difference ◽  
Km Value ◽  
Sweet Potato Starch ◽  
Methoxy Polyethylene Glycol

The sweet potato β-amylase (SPA) was modified by 6 types of methoxy polyethylene glycol to enhance its specific activity and thermal stability. The aims of the study were to select the optimum modifier, optimize the modification parameters, and further investigate the characterization of the modified SPA. The results showed that methoxy polyethylene glycol maleimide (molecular weight 5000, Mal-mPEG5000) was the optimum modifier of SPA; Under the optimal modification conditions, the specific activity of Mal-mPEG5000-SPA was 24.06% higher than that of the untreated SPA. Mal-mPEG5000-SPA was monomeric with a molecular weight of about 67 kDa by SDS-PAGE. The characteristics of Mal-mPEG5000-SPA were significantly improved. The Km value, Vmax and Ea in Mal-mPEG5000-SPA for sweet potato starch showed that Mal-mPEG5000-SPA had greater affinity for sweet potato starch and higher speed of hydrolysis than SPA. There was no significant difference of the metal ions’ effect on Mal-mPEG5000-SPA and SPA.

scholarly journals Production and Purification of IgY antibodies from egg yolk

2020 ◽  
Vol 3 (3) ◽  
pp. 192-195
Author(s):  
Sao Mai Tran Thi ◽  
Oanh Dang Thi ◽  
Huong Dang Thi ◽  
Long Le Thanh ◽  
Lan Huong Nguyen Thi ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Antibody Production ◽  
High Purity ◽  
Bovine Serum ◽  
Significant Contribution ◽  
Polyclonal Antibodies ◽  
Scientific Research ◽  
Egg Yolk ◽  
Egg Yolks

Polyclonal antibodies from vaccinated mammalian have made a significant contribution to scientific research and diagnosis. The fact that recent technologies allow the production of antibodies in egg yolks laid by hens has led to the development of an alternative to antibody production that is less dangerous to animals. The results of this study showed that it is possible to produce a large amount of IgY antibody to bovine serum albumin (BSA), content of about 8 mg/mL of egg yolk emulsion. The anti-BSA IgY antibodies with high purity and anti-BSA activity.

scholarly journals Production and Characterization of Immunoglobuline Yolk as anti antigen membrane Toxoplasma gondii

2016 ◽  
Vol 6 (2) ◽  
pp. 29
Author(s):  
Heni Puspitasari ◽  
Yuliana Praptiwi ◽  
Lucia Suwanti
Keyword(s):  
Molecular Weight ◽  
Toxoplasma Gondii ◽  
Egg Yolk ◽  
Immunoglobulin Y ◽  
Extraction And Purification ◽  
Sds Page ◽  
Protein Membrane ◽  
Peritoneal Infection ◽  
Antigen Epitope

Toxoplasma gondii is an obligate parasite intracellular which can infected human and other mammalian. Immunoglobulin Y technology  offers several  advantages better than antibody production in mammals. This research was aimed to get immunoglobulin Y from egg yolk, to prove that antibody against membrane T. gondii antigen can produced from immunoglobulin Y and to know the characterization of  immunoglobuline Y according to  molecular weight by SDS PAGE and reactivation of  antibody antigen  by Western Blott. This research devided  from many step : passase tachyzoites T. gondii into mice by peritoneal infection, cultivated  the tachyzoite from intraperitoneal fluid, preparation of  membrane antigen tachyzoite T. gondii, then  immunization laying hens with membrane antigen, extraction and purification immunoglobuline Y from egg yolk and then protein analyzed by SDS PAGE and Western Blott.  The result of this resarch showed that immunoglobulin Y from egg yolk can  produced antibody against protein membrane T. gondii. The result of analyzed profile protein immunoglobuline  Y according SDS PAGE  has molecular weight 179,8 kDa. Analyzed from Western Blott showed that immunoglobulin Y can recognize antigen epitope of  T. gondii on molecular weight 35,7 kDa and 78,8 kDa.

Structure of Myosin Subfragment-1 from Electron Microscopy and Crystallography

1988 ◽  
Vol 46 ◽  
pp. 156-157
Author(s):  
Donald A. Winkelmann
Keyword(s):  
Electron Microscopy ◽  
Molecular Weight ◽  
Actin Filaments ◽  
Light Chains ◽  
Myosin Filament ◽  
Primary Role ◽  
Heavy Chains ◽  
Myosin Subfragment ◽  
Myosin Subfragment 1 ◽  
Globular Head

The primary role of the interaction of actin and myosin is the generation of force and motion as a direct consequence of the cyclic interaction of myosin crossbridges with actin filaments. Myosin is composed of six polypeptides: two heavy chains of molecular weight 220,000 daltons and two pairs of light chains of molecular weight 17,000-23,000. The C-terminal portions of the myosin heavy chains associate to form an α-helical coiled-coil rod which is responsible for myosin filament formation. The N-terminal portion of each heavy chain associates with two different light chains to form a globular head that binds actin and hydrolyses ATP. Myosin can be fragmented by limited proteolysis into several structural and functional domains. It has recently been demonstrated using an in vitro movement assay that the globular head domain, subfragment-1, is sufficient to cause sliding movement of actin filaments.The discovery of conditions for crystallization of the myosin subfragment-1 (S1) has led to a systematic analysis of S1 structure by x-ray crystallography and electron microscopy. Image analysis of electron micrographs of thin sections of small S1 crystals has been used to determine the structure of S1 in the crystal lattice.

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