biochemical methods 2
43 Questions
5 Views

Choose a study mode

Play Quiz
Study Flashcards
Spaced Repetition
Chat to lesson

Podcast

Play an AI-generated podcast conversation about this lesson

Questions and Answers

What is the function of the stationary phase in ion exchange chromatography?

  • To bind to negatively charged molecules
  • To separate molecules based on charge or polarity (correct)
  • To serve as the mobile phase
  • To move quickly down the column

    • What type of molecule does an anion exchange resin bind to?

  • Cations
  • Negatively charged molecules
  • Anions (correct)
  • Positively charged molecules

    • When is a cation exchange resin most likely to move slowly down the column in ion exchange chromatography?

  • When negatively charged molecules are added
  • When the pH is acidic
  • When the pH is neutral
  • When positively charged molecules are added (correct)

    • What type of molecules is ion exchange chromatography generally used as a purification technique for?

    <p>Peptides, nucleotides, DNA, and proteins</p> Signup and view all the answers

    What is the net charge of a molecule?

    <p>The overall charge on the molecule when all individual charges are added together</p> Signup and view all the answers

    In what situation would an anion exchange resin move quickly down the column in ion exchange chromatography?

    <p>When positively charged molecules are added</p> Signup and view all the answers

    What is the characteristic of strong exchangers in ion exchange chromatography?

    <p>They are always ionized</p> Signup and view all the answers

    What type of molecule does a cation exchange resin bind to?

    <p>Cations</p> Signup and view all the answers

    What is the purpose of an indirect ELISA?

    <p>To detect and quantify specific proteins in a complex mixture</p> Signup and view all the answers

    In sandwich ELISA, each well of the plate is coated with:

    <p>Antibody</p> Signup and view all the answers

    What is one of the disadvantages of ELISA?

    <p>Time-consuming</p> Signup and view all the answers

    What happens if the sample in indirect ELISA contains antibodies to the antigen?

    <p>It binds to the antibodies in the well</p> Signup and view all the answers

    What type of ELISA is used for quantitating the amount of an antigen in a sample?

    <p>Sandwich ELISA</p> Signup and view all the answers

    What is added after washing the wells in sandwich ELISA?

    <p>Different antibody conjugated to an enzyme</p> Signup and view all the answers

    What does the enzyme substrate turn if antigen is present in sandwich ELISA?

    <p>Yellow</p> Signup and view all the answers

    What is used for HIV testing?

    <p>Indirect ELISA</p> Signup and view all the answers

    What does ELISA primarily offer in terms of advantages?

    <p>Specificity and sensitivity</p> Signup and view all the answers

    In ion exchange chromatography, what can be altered to elute molecules from the column?

    <p>pH or salt concentration</p> Signup and view all the answers

    What is the function of a pH gradient in protein separation using isoelectric focusing?

    <p>Separating molecules based on charge</p> Signup and view all the answers

    What is the main principle behind reverse phase chromatography?

    <p>Using a polar stationary phase and non-polar mobile phase</p> Signup and view all the answers

    What is the main advantage of TLC (thin layer chromatography)?

    <p>Visualizing components and cheap</p> Signup and view all the answers

    What is the distinguishing feature of affinity chromatography?

    <p>Separates based on affinity for a specific ligand</p> Signup and view all the answers

    What is the purpose of western blotting in conjunction with SDS-PAGE?

    <p>Visualizing specific proteins using an antibody</p> Signup and view all the answers

    What can be used to elute molecules from an ion exchange chromatography column?

    <p>pH or salt concentration</p> Signup and view all the answers

    What is the disadvantage of reverse phase chromatography?

    <p>Not good for molecules with low solubility in water</p> Signup and view all the answers

    Proteins can be separated based on charge using reverse phase chromatography.

    <p>False</p> Signup and view all the answers

    Isoelectric focusing separates molecules based on size using a pH gradient.

    <p>False</p> Signup and view all the answers

    Affinity chromatography is not suitable for all proteins.

    <p>True</p> Signup and view all the answers

    Anion exchange resin has negatively charged groups on the beads, so it binds to cations.

    <p>False</p> Signup and view all the answers

    Cation exchange resin moves slowly down the column when negatively charged molecules are added.

    <p>False</p> Signup and view all the answers

    Ion exchange chromatography is generally used as a purification technique for lipids.

    <p>False</p> Signup and view all the answers

    ELISA is a technique used to separate and quantify DNA molecules

    <p>False</p> Signup and view all the answers

    Each well of the plate in indirect ELISA is coated with an antigen

    <p>True</p> Signup and view all the answers

    Sandwich ELISA is used for quantitating the amount of antibody in a sample

    <p>False</p> Signup and view all the answers

    The enzyme substrate turns yellow if the enzyme is not bound in sandwich ELISA

    <p>False</p> Signup and view all the answers

    what does the pH needs to be for a molecule to bind to anion exchange resin?

    <p>greater than molecules isoelectric pH</p> Signup and view all the answers

    what does the Ph need to be for a molecule to bind to cation exchange resin?

    <p>less than molecules isoelectric ph</p> Signup and view all the answers

    what is the stationary phase in reverse phase chromatography?

    <p>hydrophobic beads</p> Signup and view all the answers

    what is the mobile phase in reverse phase chromatography?

    <p>polar solvent</p> Signup and view all the answers

    what does affinity chromatography beads have to attract the proteins of interest?

    <p>ligand attached to the beads</p> Signup and view all the answers

    whats the first and secondary peak for in affinity chromatography?

    <p>first= non specific proteins second= when excess affinity ligand added</p> Signup and view all the answers

    whats westeren blotting for?

    <p>find specific proteins within a mixture</p> Signup and view all the answers

    proteins are already negatively charged due to SDS-PAGE in western blotting

    <p>True</p> Signup and view all the answers

    Study Notes

    • Ion exchange chromatography: net charges of molecules change at different pH levels due to different pKa for COOH and NH2 groups.

    • To elute molecules from column, pH or salt concentration can be altered.

    • Isoelectric pH (pI): the pH at which a molecule has no net charge. Different amino acids have different pIs (e.g. aspartate at pH 3.5, lysine at pH 8.5).

    • Proteins separate based on charge using this method with a pH gradient to elute them one at a time.

    • Advantages: straightforward, good resolution, quick. Disadvantages: only works for charged molecules, some may not like pH or salt changes.

    • Isoelectric focusing: separates molecules based on charge using a pH gradient. Can be used with SDS-PAGE for size and charge separation.

    • Reverse phase chromatography: separates molecules based on polarity with hydrophobic beads and polar mobile phases. Advantages: flexibility, good resolution. Disadvantages: not good for molecules with low solubility in water.

    • TLC: thin layer chromatography separates molecules based on their interaction with a polar stationary phase and non-polar mobile phase. Advantages: can visualize components, different solvents can be used, cheap and easy. Disadvantages: small samples only, not good for proteins or large molecules.

    • Affinity chromatography: separates molecules based on their affinity for a specific ligand attached to stationary phase beads. Advantages: easy, quick, excellent purification. Disadvantages: not possible for all proteins.

    • Western blotting: visualizes specific proteins within a mixture using an antibody. Runs in conjunction with SDS-PAGE. Advantages: specific, sensitive. Disadvantages: not possible to extract protein, time-consuming.

    • Ion exchange chromatography: net charges of molecules change at different pH levels due to different pKa for COOH and NH2 groups.

    • To elute molecules from column, pH or salt concentration can be altered.

    • Isoelectric pH (pI): the pH at which a molecule has no net charge. Different amino acids have different pIs (e.g. aspartate at pH 3.5, lysine at pH 8.5).

    • Proteins separate based on charge using this method with a pH gradient to elute them one at a time.

    • Advantages: straightforward, good resolution, quick. Disadvantages: only works for charged molecules, some may not like pH or salt changes.

    • Isoelectric focusing: separates molecules based on charge using a pH gradient. Can be used with SDS-PAGE for size and charge separation.

    • Reverse phase chromatography: separates molecules based on polarity with hydrophobic beads and polar mobile phases. Advantages: flexibility, good resolution. Disadvantages: not good for molecules with low solubility in water.

    • TLC: thin layer chromatography separates molecules based on their interaction with a polar stationary phase and non-polar mobile phase. Advantages: can visualize components, different solvents can be used, cheap and easy. Disadvantages: small samples only, not good for proteins or large molecules.

    • Affinity chromatography: separates molecules based on their affinity for a specific ligand attached to stationary phase beads. Advantages: easy, quick, excellent purification. Disadvantages: not possible for all proteins.

    • Western blotting: visualizes specific proteins within a mixture using an antibody. Runs in conjunction with SDS-PAGE. Advantages: specific, sensitive. Disadvantages: not possible to extract protein, time-consuming.

    Studying That Suits You

    Use AI to generate personalized quizzes and flashcards to suit your learning preferences.

    Quiz Team

    Related Documents

    Biochemical methods 2.docx

    Description

    Learn the fundamental principles of ion exchange chromatography, a technique used to separate molecules based on their charge or polarity. Explore the process involving a mobile phase running over a stationary phase composed of charged beads, allowing the separation of molecules according to affinity.

    More Like This

    Chromatography: Distribution Constant K
    5 questions

    Chromatography: Distribution Constant K

    GraciousSynergy6241 avatar
    GraciousSynergy6241
    Chromatography: The Science of Separation
    5 questions

    Chromatography: The Science of Separation

    BetterTiger avatar
    BetterTiger
    Chromatography Applications in Organic Chemistry Quiz
    20 questions

    Chromatography Applications in Organic Chemistry Quiz

    ProgressiveSacramento avatar
    ProgressiveSacramento
    Chromatography Basics
    31 questions

    Chromatography Basics

    DynamicNarrative avatar
    DynamicNarrative
    Use Quizgecko on...
    Browser
    Open
    Browser
    Browser