Biotechnology Techniques and Chromatography Quiz

Questions and Answers

Explain the significance of the partition coefficient in chromatography.

The partition coefficient indicates the distribution of a compound between the stationary and mobile phases, which affects the separation efficiency in chromatography.

Differentiate between isopycnic and rate-zonal centrifugation.

Isopycnic centrifugation separates particles based on their density, while rate-zonal centrifugation separates based on size and buoyancy through a gradient.

What is the role of SDS in SDS-PAGE?

SDS (Sodium Dodecyl Sulfate) denatures proteins and imparts a negative charge, allowing for separation based solely on size during electrophoresis.

Describe how dialysis works for separation purposes.

Dialysis works by allowing small molecules to diffuse through a semi-permeable membrane while retaining larger molecules, achieving separation based on size. Signup and view all the answers

What are the primary applications of radioactive isotopes in biotechnology?

Radioactive isotopes are primarily used in tracer studies, medical diagnostics, and radiotherapy, providing insights and treatments in biological contexts. Signup and view all the answers

How does gas liquid chromatography (GLC) differ from high-performance liquid chromatography (HPLC)?

GLC is designed for volatile compounds in gas phase while HPLC is used for a wide range of non-volatile substances in liquid phase under high pressure. Signup and view all the answers

What safety measures should be taken when handling radioactive materials?

Safety measures include minimizing exposure, using protective equipment, and monitoring radiation levels to prevent DNA damage and mutations. Signup and view all the answers

Explain the principle behind affinity chromatography.

Affinity chromatography separates biomolecules based on their specific interactions with a ligand that is immobilized on the stationary phase. Signup and view all the answers

What is the main advantage of using a compound microscope over a simple microscope?

A compound microscope offers higher magnification due to multiple lenses, allowing for detailed observation of cells and tissues. Signup and view all the answers

How does a phase contrast microscope enhance the visibility of transparent specimens?

It converts phase shifts in light into variations in brightness, enhancing contrast for observing live cells. Signup and view all the answers

Describe the principle difference between SEM and TEM in electron microscopy.

SEM scans the surface to create detailed images, while TEM transmits electrons through the specimen to visualize internal structures. Signup and view all the answers

What is the significance of the Beer-Lambert Law in photometry?

The Beer-Lambert Law relates absorbance to the concentration of a solution, allowing for quantitative analysis. Signup and view all the answers

List two methods used for tissue homogenization and briefly describe how they work.

Motor and pestle grind tissue mechanically, while sonicators use ultrasonic waves to disrupt cells. Signup and view all the answers

In the context of protein purification, explain the difference between salting-in and salting-out.

Salting-in increases protein solubility with lower salt concentrations, whereas salting-out decreases solubility, precipitating proteins at higher salt concentrations. Signup and view all the answers

What role do chemicals play in the process of tissue homogenization?

Chemicals can specifically lyse cellular membranes, assisting in the breakdown of tissues for sample preparation. Signup and view all the answers

Why is the freeze-thaw cycle utilized in homogenization, and what is its purpose?

The freeze-thaw cycle disrupts cell membranes by expanding and contracting, facilitating the release of cellular contents. Signup and view all the answers

Study Notes

Organic Solvent Extraction and Separation Techniques

Solubility-based extraction separates components in a mixture using organic solvents.
Dialysis relies on diffusion through a semi-permeable membrane to separate small molecules from larger ones.
Reverse Dialysis enhances sample concentration by allowing smaller molecules to diffuse away from a larger, concentrated solution.
Ultrafiltration employs pressure to drive the separation of molecules based on size.
Lyophilization, or freeze-drying, preserves samples by removing moisture, allowing for long-term storage.

Chromatography

Chromatography separates substances based on various factors, primarily utilizing the partition coefficient, which describes distribution between stationary and mobile phases.
The Rf value indicates how far a substance travels relative to the solvent front.

Types of Chromatography

Paper Chromatography: Economical and straightforward for separating mixtures.
Thin Layer Chromatography (TLC): Utilizes thin adsorbent layers for separation.
Adsorption Chromatography: Functions based on the adsorption of substances to solid materials.
Ion Exchange Chromatography: Separates ions and polar molecules based on charge.
Gel Filtration Chromatography: Differentiates molecules based on size.
Affinity Chromatography: Uses specific binding interactions for targeted separation.
Gas Liquid Chromatography (GLC): Suitable for analyzing volatile compounds.
High Performance Liquid Chromatography (HPLC): Provides precise and efficient separations.

Electrophoresis

Types of Electrophoresis

Paper Electrophoresis: Simple and cost-effective method for separation.
Gel Electrophoresis: Incorporates gels for improved resolution.
PAGE (Polyacrylamide Gel Electrophoresis): Effective for separating proteins based on size.
SDS-PAGE: Denatures proteins for size-based separation.
Agarose Gel Electrophoresis: Commonly used for DNA and RNA analysis.
Isoelectric Focusing: Separates proteins based on their isoelectric points.

Centrifugation

Utilizes centrifugal force to segregate materials based on density.
Sedimentation Coefficient indicates the rate of particle sedimentation.
RPM (Revolutions per minute) quantifies centrifugal speed, while RCF reflects relative centrifugal force.

Types of Centrifuges

Swinging Bucket Rotor: Offers horizontal separation for samples.
Fixed Angle Rotor: Enables quicker sedimentation.

Centrifuge Types

Low-Speed Centrifuge: Used for general separations.
High-Speed Centrifuge: Suitable for more complex separation processes.
Ultracentrifuge: Designed for separating macromolecules at high speeds.

Centrifugation Methods

Differential Centrifugation: Involves sequential increases in speed for separation.
Density Gradient Centrifugation: Segregates samples based on density.

Radioactivity, Isotopes, and Applications

Radioisotopes are unstable isotopes that emit radiation.
Key measurement units include:
- Becquerel (Bq): One decay per second.
- Curie (Ci): 3.7 × 10^10 decays per second.
- Rutherford (Rd): 10^6 decays per second.
Disintegration Constant (λ): Measures the probability of decay.
Half-Life (t½): Time required for half the substance to decay.

Detection Techniques

Solid Scintillation Counter: Detects radiation via light emission.
Liquid Scintillation Counter: Measures beta particles in liquids.
Geiger Muller Counter: General-purpose radiation detection tool.

Applications

Tracer studies monitor substances in biological systems.
Medical diagnostics use radioactivity for imaging.
Radiotherapy treats cancer with targeted radiation.

Autoradiography

Visualizes substances utilizing radioactive labeling techniques.
Exposes samples to film, followed by development to reveal labeled areas.

Hazards and Safety

Risks include DNA damage, mutations, and cancer.
Safety measures: Minimize exposure, use protective equipment, monitor exposure levels.

Microscopy

Types of Microscopes

Simple Microscope: Utilizes a single lens for basic magnification.
Compound Microscope: Combines multiple lenses for more detailed observations.
Phase Contrast Microscope: Enhances contrast for live and transparent samples.

Electron Microscopes

SEM (Scanning Electron Microscope): Scans surfaces with electrons for detailed images.
TEM (Transmission Electron Microscope): Visualizes internal structures by passing electrons through samples.

Sample Preparation for Electron Microscopy

Involves fixation, dehydration, embedding, sectioning, and staining processes.

Photometry and Fluorimetry

Beer-Lambert Law: Connects absorbance to concentration.
Absorption spectra illustrate how substances absorb light at various wavelengths.

Instrumentation and Applications

Colorimeter: Measures absorbance at specific wavelengths.
Spectrophotometer: Evaluates absorbance over a range of wavelengths.
Fluorimeter: Measures fluorescence emitted by samples.
Flame Photometer: Analyzes metal ion concentrations in solutions.

Homogenization and Purification Methods

Tissue Homogenization Techniques

Motor and Pestle: Mechanical grinding of tissues.
Sonicator: Utilizes ultrasonic waves for tissue disruption.
Blender: Employs mechanical shearing for homogenization.
Dyno Mill: Grinds tissues using beads.
High-Pressure Homogenizer: Applies high pressure for effective breakdown.

Additional Homogenization Techniques

Osmotic Shock: Alters osmotic pressure for cell lysis.
Detergents: Solubilize cellular membranes.
Chemicals: Utilize specific agents for lysis.
Freeze-Thaw Cycle: Employs repeated freezing and thawing to disrupt cells.
Enzymes: Engages enzymes for targeted digestion.

Purification Methods

Salting-In & Salting-Out: Adjusting salt concentration influences protein solubility.

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Description

Test your knowledge on essential biotechnology techniques, including chromatography, centrifugation, and SDS-PAGE. This quiz will explore the significance of the partition coefficient, the roles of different separation methods, and the applications of radioactive isotopes in the field. Challenge yourself with these key concepts in biotechnology!