Microbiology Study Notes on E. coli and C. elegans
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Questions and Answers

What is the maximum magnification power of a conventional light microscope?

  • 150,000 X
  • 10,000 X
  • 200,000 X
  • 1,000 X (correct)

What is the resolving power of a transmission electron microscope?

  • 0.5 mm
  • 1-2 nm (correct)
  • 0.1 mm
  • 0.2 mm

Which of the following components is part of the mechanical system of a conventional optical microscope?

  • Objective lens
  • Condenser
  • Eyepiece
  • Stage (correct)

What is the typical numerical aperture for air in optical microscopy?

<p>1.0 (A)</p> Signup and view all the answers

Which type of microscopy provides the highest magnification?

<p>Transmission electron microscopy (B)</p> Signup and view all the answers

What is the resolution limit of the human eye in millimeters?

<p>0.1 mm (D)</p> Signup and view all the answers

Which system of a conventional light microscope includes the spotlight and diaphragm?

<p>Lighting system (A)</p> Signup and view all the answers

Using which type of microscope would you best observe a polypeptide chain?

<p>Transmission electron microscope (B)</p> Signup and view all the answers

What does Forward Scatter (FSC) measure in cell analysis?

<p>The relative size of the cell (C)</p> Signup and view all the answers

What is the primary purpose of breaking the plasma membrane in cellular studies?

<p>To obtain a suspension of subcellular components (A)</p> Signup and view all the answers

Which method is commonly used to break down cell membranes for cellular studies?

<p>Osmotic shock (A)</p> Signup and view all the answers

What does Side Scatter (SSC) indicate during cell analysis?

<p>The internal complexity or granularity of the cell (C)</p> Signup and view all the answers

What is differential centrifugation primarily used for?

<p>To separate macromolecules and cellular components (A)</p> Signup and view all the answers

What is the purpose of density gradient centrifugation?

<p>To separate organelles by sedimentation based on a density gradient (C)</p> Signup and view all the answers

How fast does an ultracentrifuge typically operate?

<p>100,000 rpm (D)</p> Signup and view all the answers

Which enzyme is commonly used in enzymatic methods to break down cell walls?

<p>Lysozyme (A)</p> Signup and view all the answers

What is the primary purpose of immunohistochemical techniques?

<p>To identify the presence of a specific protein in a tissue section (A)</p> Signup and view all the answers

What allows multiphoton microscopy to emit fluorescence only from the focus plane of the laser?

<p>Simultaneous absorption of two or more photons (A)</p> Signup and view all the answers

Which technique is most widely used in immunohistochemistry?

<p>Indirect method using conjugated secondary antibodies (C)</p> Signup and view all the answers

What is the theoretical maximum resolution of an electron microscope?

<p>0.002 nm (C)</p> Signup and view all the answers

What staining agents are mentioned for contrasting in electron microscopy?

<p>Uranium acetate and lead citrate (D)</p> Signup and view all the answers

Which component is NOT part of the flow cytometer?

<p>Primary antibody conjugation unit (D)</p> Signup and view all the answers

What limits the resolution of biological samples in transmission electron microscopy (TEM)?

<p>Lack of contrast due to absorption (C)</p> Signup and view all the answers

Which component modifies the electron beam before it reaches the biological sample in a TEM?

<p>Electromagnetic coils (C)</p> Signup and view all the answers

In flow cytometry, how are cells analyzed?

<p>As they pass through a laser in a fine capillary (A)</p> Signup and view all the answers

What essential factor is needed for electron microscopy that prevents in vivo studies?

<p>Maintaining a vacuum (A)</p> Signup and view all the answers

What characterizes the indirect method of immunohistochemistry?

<p>The signal is generated by a secondary antibody. (B)</p> Signup and view all the answers

What is one of the main applications of flow cytometry?

<p>Quantifying cell populations specifically labeled with fluorescent substances (A)</p> Signup and view all the answers

Which of the following is a contrast substance commonly used in TEM?

<p>Uranium acetate (B)</p> Signup and view all the answers

What is the diameter range for the thickness of samples in transmission electron microscopy?

<p>10-100 nm (A)</p> Signup and view all the answers

What is a significant advantage of using indirect immunohistochemical techniques?

<p>They provide a stronger signal with less primary antibodies. (C)</p> Signup and view all the answers

What technique is more powerful than light microscopy for analyzing details of cell structures?

<p>Electron microscopy (B)</p> Signup and view all the answers

What is the primary purpose of fluorescence microscopy?

<p>To visualize structures selectively labeled with fluorescent substances. (B)</p> Signup and view all the answers

What distinguishes green fluorescent protein (GFP) in fluorescence microscopy?

<p>It can be fused to any protein of interest using recombinant DNA technology. (C)</p> Signup and view all the answers

Which microscopy technique is most effective for observing living cells with different refractive indices?

<p>Differential interference-contrast microscopy. (B)</p> Signup and view all the answers

What role does the dichroic mirror play in fluorescence microscopy?

<p>It selects the excitation light wavelength and deflects it towards the sample. (B)</p> Signup and view all the answers

What type of cellular components can be specifically stained using fluorescent dyes?

<p>Any cellular component, including proteins and organelles. (C)</p> Signup and view all the answers

Which of the following is NOT a feature of differential interference-contrast microscopy?

<p>It produces color images of cellular components. (D)</p> Signup and view all the answers

What substance is used to stain the nucleus selectively in fluorescence microscopy?

<p>DAPI. (D)</p> Signup and view all the answers

What is the effect of MitoTracker Red CMXRos on cell observation?

<p>It labels the mitochondrial network within cells. (B)</p> Signup and view all the answers

What is the primary difference between speed separations and equilibrium separations?

<p>Speed separations depend on particle size, while equilibrium separations depend on particle density. (A)</p> Signup and view all the answers

What role do gradients such as Ficoll, Percol, or Nicodenz play in centrifugation?

<p>They create a concentration gradient for separating cells based on density. (B)</p> Signup and view all the answers

Which statement accurately describes the process of cell culture?

<p>Cell culture requires rigorous control of experimental conditions. (B)</p> Signup and view all the answers

What defines a clone in cell culture?

<p>A genetically homogeneous strain of cells derived from one original cell. (C)</p> Signup and view all the answers

How do primary cultures differ from secondary cultures?

<p>Primary cultures are established from a larger tissue sample, while secondary cultures are derived from a primary culture. (C)</p> Signup and view all the answers

Why are tumor-derived cells referred to as immortal cells?

<p>They can grow indefinitely in culture. (D)</p> Signup and view all the answers

What milestone in cell biology was achieved in 1951?

<p>The first immortal human cell line was obtained. (B)</p> Signup and view all the answers

What underlies the process of separating Peripheral Blood Mononuclear Cells (PBMC)?

<p>They are isolated based on their density using specific gradients. (B)</p> Signup and view all the answers

Flashcards

Resolving power

The ability of a microscope to distinguish two closely spaced objects as separate entities.

Magnification

Ability of the objective lens to make an image appear larger than it actually is.

Light microscopy

A type of microscopy that uses light to illuminate and magnify a sample.

Electron microscopy

A type of microscopy that uses electrons to illuminate and magnify a sample.

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Transmission electron microscopy (TEM)

A type of electron microscopy that transmits electrons through a thin sample, producing a high-resolution image.

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Scanning electron microscopy (SEM)

A type of microscopy that illuminates a sample from the side, producing a 3D image of surface details.

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Fluorescence microscopy

A type of microscopy that uses fluorescent dyes to illuminate specific structures within a cell.

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Super-resolution microscopy

A type of microscopy that allows you to see structures smaller than the diffraction limit of light.

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Flow Cytometry

A technique that uses a laser to count, analyze the size, and evaluate the internal complexity of cells.

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Forward Scatter (FSC)

Measures the relative size of a cell by detecting light scattering in the forward direction.

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Side Scatter (SSC)

Measures the internal complexity or granularity of a cell by detecting light scattering at a 90° angle.

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Cell Disruption

The process of breaking down cells to isolate specific organelles or other subcellular components.

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Differential Centrifugation

A technique that separates subcellular components based on their size and density by spinning them at high speed.

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Density Gradient Centrifugation

A method that uses a gradient of density to separate subcellular components based on their buoyant density.

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Ultrasound

A physical method of cell disruption that uses high-frequency sound waves to break down cell membranes.

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Lysozyme

An enzyme that breaks down bacterial cell walls, often used in cell disruption.

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Multiphoton Microscopy

A microscopic technique that uses two or more photons to excite a fluorescent dye, resulting in fluorescence only from the focal plane, allowing for deeper imaging than traditional microscopy.

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Heavy Metal Staining in TEM

A method for increasing contrast in TEM images by adding heavy metals to the sample, which bind to specific structures and absorb electrons differently.

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Resolution

The distance between two points that can be distinguished as separate entities in an image.

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Numerical Aperture (NA)

A measure of the angle of light that can enter a lens, influencing the resolution of the image.

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Resolution Limit

The minimum distance between two objects that can be distinguished as separate entities.

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Confocal Microscopy

The use of a focused laser beam to excite fluorescent molecules in a sample, creating an image of the sample.

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Speed Separation

Separating particles by size. Larger particles settle faster, forming distinct bands.

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Equilibrium Separation

Separating particles by density. Particles settle until they reach their equilibrium position where their density matches the medium.

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Cell Culture

A nutrient-rich medium designed to support cell growth and survival outside of a living organism. It's like a 'cell spa'.

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Primary Culture

The initial population of cells grown from a tissue sample. Think of them as the 'first generation' of cells.

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Immortal Cells

Cells that have been repeatedly sub-cultured and can divide indefinitely. These cells are like 'super-cells'.

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Embryonic Stem Cells

Cells that can develop into different cell types. Think of them as 'master cells'.

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Differentiated Cells

Cells that can only develop into a specific cell type. Think of them as 'specialized cells'.

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Cloned Cells

Cells that have been grown from a single cell and are genetically identical. They're like 'clones'.

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What is the principle behind light microscopy staining?

A staining technique that uses a basic dye (hematoxylin) and an acidic dye (eosin) to differentiate acidic and basic structures within cells.

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How is contrast achieved in Electron Microscopy?

Electron microscopy uses high molecular weight molecules like uranium acetate and lead citrate to enhance the contrast and visibility of samples.

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What is Immunohistochemistry?

Immunohistochemistry employs specific antibodies to identify the presence of a particular protein within a tissue section.

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What is Direct Immunohistochemistry?

Direct Immunohistochemistry directly labels the primary antibody with a detectable substance, like an enzyme, fluorochrome, or gold particle.

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What is Indirect Immunohistochemistry?

Indirect Immunohistochemistry uses a secondary antibody that recognizes the primary antibody, and the secondary antibody is labeled with a detectable substance.

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What is Flow Cytometry?

Flow cytometry is a technique that analyzes the number, size, and complexity of cells within a suspension. It also quantifies labeled cells and isolates specific populations.

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Describe the components of a flow cytometer.

Flow cytometry uses a fluidic system to transport cells, a laser to illuminate them, and electronic detectors to convert light into digital signals.

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How do cells get analyzed in a flow cytometer?

Cells pass through a narrow capillary in a flow cytometer, where they are individually illuminated by a laser beam. The scattered light pattern is then detected and analyzed.

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Differential Interference Contrast (DIC) Microscopy

A microscopy technique that uses the difference in refractive index between different components within a cell to create an image. This technique is useful for visualizing living cells and their internal structures.

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DAPI (4',6-diamidino-2-phenylindole)

A commonly used fluorescent dye that selectively stains DNA. It is often used in fluorescence microscopy to visualize the nucleus of a cell.

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Green Fluorescent Protein (GFP)

A naturally occurring fluorescent protein isolated from the jellyfish Aequorea victoria. It can be genetically fused to other proteins, allowing scientists to track the movement and localization of specific proteins within cells.

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Phase Contrast Microscopy

A microscopy technique that uses a special condenser and an objective lens to enhance the contrast of unstained, transparent specimens. It enhances the visualization of phase differences in the sample.

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Phalloidin

A fluorescent dye that binds to filamentous actin, a protein involved in cell structure and movement. It is often used in fluorescence microscopy to visualize the cytoskeleton of cells.

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MitoTracker Red CMXRos

A fluorescent dye that specifically stains mitochondria, allowing scientists to visualize the mitochondrial network within cells.

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Study Notes

E. coli Study Notes

  • Under optimal conditions, E. coli divides every 20 minutes.
  • Rapid growth requires glucose, salts, and organic compounds like amino acids, vitamins, and nucleic acid precursors.
  • Slow growth requires salts, a nitrogen source (e.g., ammonia), and a carbon and energy source (e.g., glucose).

Caenorhabditis elegans Study Notes

  • It is a multicellular organism shaped like an elongated tube, thinning at the ends.
  • It has 959 somatic cells and 1000-2000 germ cells.
  • It is easily reproduced and genetically manipulated in labs.
  • Its body is covered in a thin outer cuticle.
  • Its body is organized into simple organs and systems, including a digestive system (stoma, pharynx, intestine), sexual organs (gonads), and a rudimentary nervous system.

C. elegans Genome Study Notes

  • It was the first multicellular organism to have its genome sequenced (1998).
  • Its genome is about 100 million base pairs long and has six chromosomes plus one mitochondrial chromosome.
  • It has an estimated 19,000 protein-coding genes.
  • About 36% of its genes have human homologues.

Danio rerio Study Notes

  • Zebrafish (Danio rerio) are small, active fish common in aquariums.
  • Their natural habitat includes calm, sometimes stagnant waters of central Asia, particularly the Ganges region of India.
  • Adults are typically 3-5 cm long and 1 cm wide, depending on environmental conditions.
  • They have an elongated body shape with a dorsal fin and 5-9 bluish bands that run along their sides.
  • Males are golden, while females are silver.
  • Their ventral area is whitish/pink, earning them the name "zebrafish."

Stem Cell Organoid Cultures Study Notes

  • Various tissues can be cultured using stem cells: intestine, lung, brain, stomach, pancreas, etc.
  • Activin A, BMP4, Wnt, FGF, and other growth factors are involved in organoid development.
  • ECM embedding is a key component of culture formation.

Organoid Models Study Notes

  • 2D and 3D organoid culture models are used.
  • 3D models can be better than 2D models, except for some scenarios.
  • Animal models are also used for some organoids.

Microscopy Techniques Study Notes

  • Optical microscopy types include light field, phase contrast, differential interference contrast (DIC), fluorescence, confocal, and multiphoton.
  • Light microscopy - uses visible light to magnify image.
  • Phase contrast microscopy - manipulates light to reveal internal structures that are virtually invisible in normal light microscopy.
  • DIC microscopy - a contrast enhancing technique that illuminates images with polarized light.
  • Fluorescence microscopy - uses fluorescent dyes or proteins to detect specific cellular components.
  • Confocal microscopy - employs laser light to focus on a defined spot, eliminating out-of-focus light through pinholes, creating 3D images.
  • Multiphoton microscopy - requires multiple photons for excitation of fluorophores, creating highly detailed images with minimal light scatter.
  • Electron microscopy (TEM/SEM) - uses electrons rather than light waves, magnifying images much more. TEM magnifies thin slices of specimen while SEM illuminates whole specimens.
    • Specimen preparation for light microscopy might use fixing, dehydration, clearing, infiltration, and embedding techniques.
  • Staining techniques use dye to enhance contrast.

Flow Cytometry Study Notes

  • Flow cytometry analyzes the number, size, and intracellular complexity of cells in a suspension.
  • It uses a laser to illuminate cells, allowing for analysis of forward and 90-degree scatter to determine size and complexity/granularity, respectively.
  • Flow cytometry can be used to isolate specific cell populations.
  • Subcellular components can be separated, purified, and analyzed.

Cell Cultures Study Notes

  • The study of cells in a living organism is complex.
  • Cells grown in culture can be isolated, survive, and grow under controlled conditions.
  • Cultured cells have advantages for research including controlled conditions and ease of observation.

Viruses Study Notes

  • Viruses need a host cell to carry out their life cycle.
  • Viruses can be studied in host cell cultures.
  • Using viruses, scientists can learn more about fundamental cell biology including molecular mechanisms, RNA genes, and oncogene discovery.

Immunohistochemical Techniques Study Notes

  • Immunohistochemistry is a technique used to identify specific proteins within tissue sections through the use of antibodies.
  • It can be used to analyze diseases, specific proteins, and oncogene overexpression.
  • Antibody labeling methods can be direct (primary antibody labeled) or indirect (primary antibody detected by labeled secondary antibody).

Cell Biology Instruments Notes

  • Specimen preparation, flow cytometry, subcellular separation, and growth of animal cells are common instruments in cell biology.
  • Histological processing (fixation, tissue embedding, sectioning, and staining) is vital for preparing specimen for analysis.
  • Specific techniques, like differential centrifugation and density gradient centrifugation, are important for purifying specific organelles and other components within cells.

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Description

Explore the fascinating world of E. coli and Caenorhabditis elegans through these study notes. Learn about their growth conditions, cellular structures, and genetic traits, including C. elegans being the first multicellular organism to have its genome sequenced. This summary captures essential information for microbiology enthusiasts.

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