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plasma proteins

A
Amna Sheikh

The document discusses plasma proteins, including their definition, functions, compositions, detection methods, and electrophoresis. Plasma proteins comprise 7% of plasma solids and include albumin, globulins, fibrinogen, and others. They serve important roles like nutrient transport, coagulation, and osmotic balance. Detection methods discussed are precipitation, electrophoresis, immunoassays, and more. Serum protein electrophoresis separates proteins by size and charge and is commonly used to analyze plasma protein composition.

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PLASMA PROTEINS Chemical pathology Presented by Amna Sahar
Contents Definition of plasma proteins Functions Compositions Follow-up plasma protein test Methods for detection of different plasma proteins Protein electrophoresis References
Plasma Proteins The proteins present in the plasma of human blood are a mixture of simple proteins, glycoproteins, lipoproteins and other conjugated proteins are called Plasma Proteins. These may be separated by salt precipitation, immunological technique and electrophoresis Plasma is obtained from anti-coagulated blood Plasma proteins forms 7% of the solids in plasma Total protein content of normal plasma is 6-8g/100ml Almost all plasma proteins are synthesized by liver except immunoglobulin
Functions Protein nutrition Osmotic pressure and water balance Buffering action Transport of lipids Transport of other substances Blood coagulation
Composition of Plasma Proteins Plasma proteins composed of : Albumin 55.2% Globulin 留1-globulin 5.3% 留2-globulin 8.6% -globulin 13.4% 粒-globulin 11.0 % Fibrinogen 6.5%
Further Composition of Plasma Proteins Retinol binding protein 留1-fetoprotein (AFP) 留1-antitrypsin 留1-protease inhibitor (API) 留1-acid glycoprotein (AAG) High density lipoprotein (HDL) Prothrombin 留1-globulin 5.3%
Further Composition of Plasma Proteins Ceruloplasmin (ferro-oxidase) Corticosteroid binding globulin Hepatoglobin Thyroxin binding globulin (TBG) 留2-macroglobulin (AMG) 留2-globulin 5.3%
Further Composition of Plasma Proteins Transferrin Hemopexin Low density lipoprotein LDL 硫2 macroglobulin C4 complement C3 compliment C1q complement C-reactive protein 硫-globulin 13.4%
Further Composition of Plasma Proteins Ig G Ig M Ig E Ig A Ig D 粒-globulin 11.0%
Follow Up Plasma Protein Test C-reactive protein tests to evaluate for inflammation Immunoglobulin tests to measure antibodies liver enzyme tests Protein electrophoresis to look for underlying bone marrow disorders
Methods for detection of Plasma Protein Precipitation method BCG method electrophoresis Ultra centrifugation Precipitation method Radioimmunoassay Enzyme-labelled Immunoassay Serum protein electrophoresis Albumin For 留1-fetoprotein (AFP) 留1-Globulin
Methods for detection of Plasma Protein Precipitation method Serum protein electrophoresis Immunodiffusion Immunonephelometric assay Immunofixation Radial immunodiffusion Immunoturbidity Nephelometry Immunofixation For 留1-anti trypsin For 留1-acid glycoprotein
Methods for detection of Plasma Protein Radial immunodiffusion Immunonephelometric Radial immunodiffusion Immunonephelometric For Hepatoglobin 留2-Globulin For ceruloplasmin Radial immunodiffusion Immunonephelometric ELISA Latex agglutination immunoassay For 留2-microglobin
Methods for detection of Plasma Protein Radial immunodiffusion Immunonephelometric TIBC Radial immunodiffusion ELISA Nephelometry CRP assay For Transferrin 硫-Globulin For Hemopexin Immunoassay Serum protein electrophoresis Precipitation Nephelometric immunoassay Turbidimetry For 硫2-microglobin For LDL For ComplementCRP
Methods for detection of Plasma Protein Radial immunodiffusion Nephelometry Turbidimetry Electro chemiluminescent immunoassay Radioimmunoassay Screening Serum protein electrophoresis 粒-Globulin
Methods for detection of Plasma Protein Radial immunodiffusion Nephelometry Turbidimetry Radioimmunoassay Plasma protein electrophoresis Clotting time CT Fibrinogen
Principles of Methods for detection of Plasma Protein Radial immunodiffusion An antigen sample is placed in a well and allowed to diffuse into agar containing a suitable dilution of an antiserum. As the antigen diffuses into the agar, the region of equivalence is established and a ring of precipitation, a precipitin ring, forms around the well. The area of the precipitin ring is proportional to the concentration of antigen. . Nephelometry The principle is to measure forwarded scattered light when a laser beam passes through a sample and the light is deflected by the particles. Radioimmunoassay The principle of RIA involves competitive binding of radiolabeled antigen and unlabeled antigen to a high-affinity antibody. The labeled antigen is mixed with antibody at a concentration that saturates the antigen-binding sites of the antibody. Then test samples of unlabeled antigen of unknown concentration are added in progressively larger amounts The more unlabeled antigen is present, the less radioactivity there is in the complex. The concentration of the unknown (unlabeled) antigen or hapten is determined by comparison with the effect of standards.
Plasma Protein Electrophoresis Electrophoresis is a separations technique that is based on the mobility of ions in an electric field. Ions have different migration rates depending on their total charge, size, and shape, and can therefore be separated. The technique is used particularly for macromolecules, such as proteins. Gel electrophoresis involves the use of gel as supporting media for separation of DNA, RNA or proteins under the influence of electric charge. It is usually performed for analytical purposes but may be used as a preparative technique to partially purify molecules prior to use for other methods such as mass spectrometry, PCR, cloning, DNA sequencing and immuno-blotting. This is the most commonly used electrophoresis in biotechnology laboratories
Plasma Protein Electrophoresis It may be of two types; 1. Agarose gel electrophoresis Mostly for the separation of DNA fragments having more than 50 base pairs 2. PAGE (polyacrylamide gel electrophoresis) For separation of proteins It may be Sodium Dodecyl Sulfate SDS-PAGE and NATIVE
Sodium Dodecyl Sulfate (SDS) polyacrylamide gel electrophoresis is mostly used to separate proteins accordingly by size. This is one of the most powerful techniques to separate proteins on the basis of their molecular weight. Principle This technique uses anionic detergent Sodium Dodecyl Sulfate (SDS) which dissociates proteins into their individual polypeptide subunits and gives a uniform negative charge along each denatured polypeptide. Power Supply: A power supply of 100-200 volts is needed. This is ideal for running and transferring protein resolving gels. Buffer: Two types of buffers are used in SDS-PAGE. The lower reservoir (which has the running gel) has amine buffers. It is adjusted by using HCl. The upper reservoir (which has stacking gel) also has amine buffers but its pH is slightly above that of running gel buffer and is adjusted with glycine instead of HCl.Stain Coomassie Brilliant Blue R-250 (CBB) is the most popular protein stain (blue bands). 2- SDS-PAGE Reader Densitometric scanning machine
Procedure
plasma proteins
References Books Bishop Tietz Pictures https://www.biochemden.com/plasma-proteins/

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plasma proteins

  • 2. Contents Definition of plasma proteins Functions Compositions Follow-up plasma protein test Methods for detection of different plasma proteins Protein electrophoresis References
  • 3. Plasma Proteins The proteins present in the plasma of human blood are a mixture of simple proteins, glycoproteins, lipoproteins and other conjugated proteins are called Plasma Proteins. These may be separated by salt precipitation, immunological technique and electrophoresis Plasma is obtained from anti-coagulated blood Plasma proteins forms 7% of the solids in plasma Total protein content of normal plasma is 6-8g/100ml Almost all plasma proteins are synthesized by liver except immunoglobulin
  • 4. Functions Protein nutrition Osmotic pressure and water balance Buffering action Transport of lipids Transport of other substances Blood coagulation
  • 5. Composition of Plasma Proteins Plasma proteins composed of : Albumin 55.2% Globulin 留1-globulin 5.3% 留2-globulin 8.6% -globulin 13.4% 粒-globulin 11.0 % Fibrinogen 6.5%
  • 6. Further Composition of Plasma Proteins Retinol binding protein 留1-fetoprotein (AFP) 留1-antitrypsin 留1-protease inhibitor (API) 留1-acid glycoprotein (AAG) High density lipoprotein (HDL) Prothrombin 留1-globulin 5.3%
  • 7. Further Composition of Plasma Proteins Ceruloplasmin (ferro-oxidase) Corticosteroid binding globulin Hepatoglobin Thyroxin binding globulin (TBG) 留2-macroglobulin (AMG) 留2-globulin 5.3%
  • 8. Further Composition of Plasma Proteins Transferrin Hemopexin Low density lipoprotein LDL 硫2 macroglobulin C4 complement C3 compliment C1q complement C-reactive protein 硫-globulin 13.4%
  • 9. Further Composition of Plasma Proteins Ig G Ig M Ig E Ig A Ig D 粒-globulin 11.0%
  • 10. Follow Up Plasma Protein Test C-reactive protein tests to evaluate for inflammation Immunoglobulin tests to measure antibodies liver enzyme tests Protein electrophoresis to look for underlying bone marrow disorders
  • 11. Methods for detection of Plasma Protein Precipitation method BCG method electrophoresis Ultra centrifugation Precipitation method Radioimmunoassay Enzyme-labelled Immunoassay Serum protein electrophoresis Albumin For 留1-fetoprotein (AFP) 留1-Globulin
  • 12. Methods for detection of Plasma Protein Precipitation method Serum protein electrophoresis Immunodiffusion Immunonephelometric assay Immunofixation Radial immunodiffusion Immunoturbidity Nephelometry Immunofixation For 留1-anti trypsin For 留1-acid glycoprotein
  • 13. Methods for detection of Plasma Protein Radial immunodiffusion Immunonephelometric Radial immunodiffusion Immunonephelometric For Hepatoglobin 留2-Globulin For ceruloplasmin Radial immunodiffusion Immunonephelometric ELISA Latex agglutination immunoassay For 留2-microglobin
  • 14. Methods for detection of Plasma Protein Radial immunodiffusion Immunonephelometric TIBC Radial immunodiffusion ELISA Nephelometry CRP assay For Transferrin 硫-Globulin For Hemopexin Immunoassay Serum protein electrophoresis Precipitation Nephelometric immunoassay Turbidimetry For 硫2-microglobin For LDL For ComplementCRP
  • 15. Methods for detection of Plasma Protein Radial immunodiffusion Nephelometry Turbidimetry Electro chemiluminescent immunoassay Radioimmunoassay Screening Serum protein electrophoresis 粒-Globulin
  • 16. Methods for detection of Plasma Protein Radial immunodiffusion Nephelometry Turbidimetry Radioimmunoassay Plasma protein electrophoresis Clotting time CT Fibrinogen
  • 17. Principles of Methods for detection of Plasma Protein Radial immunodiffusion An antigen sample is placed in a well and allowed to diffuse into agar containing a suitable dilution of an antiserum. As the antigen diffuses into the agar, the region of equivalence is established and a ring of precipitation, a precipitin ring, forms around the well. The area of the precipitin ring is proportional to the concentration of antigen. . Nephelometry The principle is to measure forwarded scattered light when a laser beam passes through a sample and the light is deflected by the particles. Radioimmunoassay The principle of RIA involves competitive binding of radiolabeled antigen and unlabeled antigen to a high-affinity antibody. The labeled antigen is mixed with antibody at a concentration that saturates the antigen-binding sites of the antibody. Then test samples of unlabeled antigen of unknown concentration are added in progressively larger amounts The more unlabeled antigen is present, the less radioactivity there is in the complex. The concentration of the unknown (unlabeled) antigen or hapten is determined by comparison with the effect of standards.
  • 18. Plasma Protein Electrophoresis Electrophoresis is a separations technique that is based on the mobility of ions in an electric field. Ions have different migration rates depending on their total charge, size, and shape, and can therefore be separated. The technique is used particularly for macromolecules, such as proteins. Gel electrophoresis involves the use of gel as supporting media for separation of DNA, RNA or proteins under the influence of electric charge. It is usually performed for analytical purposes but may be used as a preparative technique to partially purify molecules prior to use for other methods such as mass spectrometry, PCR, cloning, DNA sequencing and immuno-blotting. This is the most commonly used electrophoresis in biotechnology laboratories
  • 19. Plasma Protein Electrophoresis It may be of two types; 1. Agarose gel electrophoresis Mostly for the separation of DNA fragments having more than 50 base pairs 2. PAGE (polyacrylamide gel electrophoresis) For separation of proteins It may be Sodium Dodecyl Sulfate SDS-PAGE and NATIVE
  • 20. Sodium Dodecyl Sulfate (SDS) polyacrylamide gel electrophoresis is mostly used to separate proteins accordingly by size. This is one of the most powerful techniques to separate proteins on the basis of their molecular weight. Principle This technique uses anionic detergent Sodium Dodecyl Sulfate (SDS) which dissociates proteins into their individual polypeptide subunits and gives a uniform negative charge along each denatured polypeptide. Power Supply: A power supply of 100-200 volts is needed. This is ideal for running and transferring protein resolving gels. Buffer: Two types of buffers are used in SDS-PAGE. The lower reservoir (which has the running gel) has amine buffers. It is adjusted by using HCl. The upper reservoir (which has stacking gel) also has amine buffers but its pH is slightly above that of running gel buffer and is adjusted with glycine instead of HCl.Stain Coomassie Brilliant Blue R-250 (CBB) is the most popular protein stain (blue bands). 2- SDS-PAGE Reader Densitometric scanning machine
  • 23. References Books Bishop Tietz Pictures https://www.biochemden.com/plasma-proteins/