Microscopes Overview

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Questions and Answers

What does the retention factor (Rp) quantify in chromatography?

The distance solvent traveled

The concentration of the solute in the solvent

The distance traveled by a specific particle

The ratio of distances traveled by a particle and solvent (correct)

What defines the lag phase in bacterial growth?

Cells adapting to their growth conditions (correct)

Cells dying due to lack of nutrients

Cells entering a dormant state

Rapid increase in cell numbers

In fluorescence lifetime imaging microscopy (FLIM), what is primarily measured?

The movement of molecules within a cell

The lifetime of emitted fluorescence from cells (correct)

The intensity of light emitted by the cells

The rate of photobleaching in cells

Which of the following describes the shape of coccus bacteria?

Spherical Signup and view all the answers

Which mechanism allows bacteria to acquire genetic material from their environment?

Transformation Signup and view all the answers

What is the main limitation of an optical microscope compared to an electron microscope?

It has low resolution. Signup and view all the answers

Which steps are involved in cell fractionation?

Extraction, homogenization, and centrifugation. Signup and view all the answers

Which organelle is likely to be found at the bottom of a density centrifugation experiment?

Nucleus Signup and view all the answers

What is the primary purpose of karyotyping?

To count chromosomes for abnormalities. Signup and view all the answers

What is one of the primary methods for determining the order of nucleotides in DNA?

DNA Sequencing. Signup and view all the answers

Which technique is used to directly find the gene causing a disease?

Single Nucleotide Polymorphisms (SNP). Signup and view all the answers

Which of the following is NOT a step in the Polymerase Chain Reaction (PCR) process?

Prioritization. Signup and view all the answers

What is the role of restriction enzymes in recombinant DNA technology?

To cut DNA at specific sequences. Signup and view all the answers

What is the function of Tag Polymerase during elongation at 70°C?

It adds nucleotides to the 3' ends. Signup and view all the answers

Which technique is used to detect specific pieces of proteins?

Western Blotting Signup and view all the answers

What is the result of applying SDS in proteomics?

It denatures proteins and adds charges. Signup and view all the answers

How does gel electrophoresis separate macromolecules?

By size and charge. Signup and view all the answers

What does genomic library store?

The DNA of an organism's genome. Signup and view all the answers

In which cell type are eukaryotic genes typically injected for bacterial cloning?

Prokaryotic cells Signup and view all the answers

What do DNA microarrays measure?

Gene expression levels. Signup and view all the answers

Which type of cell can evolve into a complete organism?

Totipotent cells Signup and view all the answers

What is the primary purpose of fixation in microscopy?

To preserve the shape and adhere specimens to the slide Signup and view all the answers

Which microscope is specifically designed to view the surface of a sample that has been contaminated or dehydrated?

Scanning Electron Microscopy (SEM) Signup and view all the answers

What is a significant feature of the Phase Contrast Microscope?

It provides high contrast through phase shifts and a Halo Effect. Signup and view all the answers

What is the result of staining a specimen in microscopy?

It enhances the visibility of the specimen. Signup and view all the answers

Which technique uses a focused laser beam to reduce artifacts in imaging?

Confocal Laser Scanning Microscopy Signup and view all the answers

In what form of microscopy is the final image characterized as a bright image against a dark background?

Dark Field Microscopy Signup and view all the answers

What additional component does a Bright Field Microscope include compared to a Compound Microscope?

An additional bright light Signup and view all the answers

Which type of microscopy provides a clear 2-D depiction of the internal structures of a cell?

Transmission Electron Microscopy (TEM) Signup and view all the answers

Study Notes

Microscopes

Fixation preserves the shape of a sample and adheres it to a slide.
Staining increases the visibility of a specimen by adding color.

Types of Microscopes

Stereo Microscope (Optical) views the exterior of a specimen, has low magnification, and creates a 2D image; used for dissections.
Compound Microscope (Optical) views simple, single-layered live cells with multiple lenses. Staining and fixation are necessary for better contrast.
Bright Field Microscope (Optical) views simple, single-layered live cells; it's identical to a compound microscope but with an additional bright light source.
Phase Contrast Microscope (Optical) views live cells that are arranged to be flat and thin, creates a 2D image, and utilizes an annular ring that refracts light to enhance contrast. Produces high phase shifts and the "Halo Effect".
Fluorescence Microscope (Optical) views various parts of a cell by exposing the specimen to ultraviolet, blue, or violet light. Uses a dichroic mirror that allows only specific wavelengths to pass, leading to clarity reduction due to distortions and artifacts.
Confocal Laser Scanning Microscopy (Optical) views fluorescent objects with the help of a laser beam focused directly on a sample to reduce artifacts. Requires longer illumination due to the laser’s focused intensity.
Dark Field Microscopy (Optical) views live, unstained cells and generates an image using light refracted by the sample. Creates a bright image against a dark background, providing high contrast.
Scanning Electron Microscopy (Electron) views the surface of a dehydrated and heavy-metal-stained sample. Produces a clear 3D depiction of the sample. (SEM)
Cryo-scanning Electron Microscopy (Electron) views the surface of a sample frozen in liquid nitrogen. Produces a clear 3D depiction of the sample. (Cryo-SEM) This is an expensive technique that introduces artifacts.
Transmission Electron Microscopy (Electron) views the interior of a cell, creating a clear 2D image of the sample. Electron tomography can be used to combine TEM images to create a 3D representation of internal structures.

Optical Microscope vs Electron Microscope

Optical Microscopes are used to view both living and non-living cells, have a low resolution, and show larger organelles like the nucleus, cytoplasm, chloroplast, and cell wall. They generate images directly.
Electron Microscopes are for non-living cells, have high resolution, and show smaller organelles like the nucleolus, ribosomes, RER, SER, centrioles, mitochondria, and lysosomes. They generate images indirectly by utilizing electrons that are passed through a magnetic field.

Biological Lab Technique

Counting Cells

Hemocytometer counts cells within a gridded area.
Colony Forming Units estimate the number of cells in a colony. (ic=10)
Automated Cell Counting utilizes a laser to detect cell numbers.

Cell Fractionation

Separates cellular components based on size and density.
The bottom of the pellet consists of larger and denser organelles compared to the top.
It involves three steps: Extraction, homogenization, and centrifugation.

Centrifugation

Separates particles based on size, shape, and density.
Differential Centrifugation separates by size, with larger particles (nucleus) sedimenting faster than smaller particles (ribosomes).
Density Centrifugation separates by density, with the bottom layer the densest and the uppermost layer the least dense.

Order of Elution / First to Last / Most Dense to Least

Nucleus
Chloroplast
Mitochondria
Lysosome
Peroxisome
Microsomes
Ribosomes

Karyotyping

Counts chromosomes using a light microscope during metaphase of mitosis to detect abnormalities.
Can diagnose Down syndrome, Turner's syndrome, and Klinefelter's syndrome.

DNA Sequencing

Determines the order of nucleotides in a DNA sequence.
Two common methods exist: older Sanger sequencing and newer Next Generation Sequencing.

Single Nucleotide Polymorphisms (SNP)

Identifies variations in the genome sequence that are used to directly locate genes responsible for specific diseases.

Recombinant DNA

Utilizes restriction enzymes to cut DNA at palindromic sequences.
Sticky ends are unpaired nucleotides, whereas blunt ends consist of paired nucleotides.
Restriction Fragment Length Polymorphisms (RFLP) is a technique that compares and contrasts DNA between individuals by examining different sized DNA fragments.

DNA Fingerprinting

Short tandem repeats and RFLP are used to identify individuals.

Polymerase Chain Reaction (PCR)

Creates millions of copies of a specific DNA segment in three steps:
- Denaturation (95°C): Heat separates DNA strands.
- Annealing (65°C): DNA primers attach to template DNA.
- Elongation (70°C): Taq Polymerase (DNA polymerase) adds nucleotides to the 3’ ends of primers.
Calculation: The number of DNA copies after "n" cycles is 2n2^n2n. The number of strands after "x" cycles is 2x22^{x2}2x2.

Bacterial Cloning

Involves injecting eukaryotic genes into prokaryotic cells.

Reverse Transcriptase

Used to create cDNA (complementary DNA) that can be inserted into bacteria using plasmids.

Plasmid

Small circular DNA molecule capable of independent replication within a bacterial cell.
Carries antibiotic resistance genes.

Gel Electrophoresis

Separates macromolecules (DNA, RNA, proteins) based on size and charge.
The top of the gel is negatively charged, while the bottom is positively charged.
Negative samples migrate towards the positive pole (+)
Positive samples migrate towards the negative pole (-)

Southern Blotting

Detects specific DNA fragments.

Northern Blotting

Detects specific RNA fragments.

Western Blotting

Detects specific protein fragments.
Utilizes primary and secondary antibodies.

ELISA (Enzyme Linked Immunosorbent Assay)

Detects the presence of specific antigens in a sample.

Pulse Chase Experiments

Tracks the movement of proteins through a cell.
- RER > GA > Secretory Vesicles > Plasma Membrane
- Pulse: Radioactive amino acids are used to label proteins.
- Chase: Prevents the division of labeled proteins.

Hershey-Chase Experiment

Labeled proteins with radioactive sulfur and DNA with radioactive phosphorus.
Showed that DNA is the transforming molecule.

Genome Annotation

Identifies the location of genes within a genome, differentiating between coding and non-coding regions.

SDS (Sodium Dodecyl Sulfate)

Denatures proteins by breaking non-covalent bonds, adding charges, and detangling them.

SDS Page

Separates proteins based on mass.

Genomics

The study of genes and their interactions within a genome.

Genomic Library

A collection of DNA fragments representing an organism's complete genome.

DNA Microarrays

Determine which genes are actively expressed, revealing the cell's transcriptome.

Transgenic Animals

An animal with a foreign gene introduced into its genome to study the gene's function.

Reproductive Cloning

Uses a somatic cell to produce a genetically identical copy of an organism.
Example: Dolly the sheep.

Totipotent

Cells capable of developing into a complete organism.

Pluripotent

Cells that can differentiate into any of the three germ layers: ectoderm, mesoderm, or endoderm.

Multipotent

Cells that can differentiate into one or two germ layers but not all three. More differentiated cells only produce one type of cell.

Chromatography

Separates components of a mixture based on their solubility.

Retention Factor (Rp)

Rp=D1D2 Rp = \frac{D_{1}}{D_{2}} Rp=D2D1
- D1D_{1}D1 = Distance of the component.
- D2D_{2}D2 = Distance of the solvent.

Fluorescence Recovery After Photobleaching (FRAP)

Cells are illuminated with a powerful laser beam, which causes a specific area to lose fluorescence.
Monitors the movement of molecules in a cell by observing the recovery of fluorescence in the bleached area.

Fluorescence Lifetime Imaging Microscopy (FLIM)

Measures the lifetime of fluorescence in a sample.
Determines the concentration of various molecules by analyzing their fluorescence lifetime.

Knockout Mouse

A genetically engineered mouse with a specific gene deleted to study its function.

Bacteria

Diplo: Pairs of bacteria.
Strep: Chains of bacteria.
Staph: Grape-like clusters of bacteria.

Coccus

Spherical bacteria.

Bacillus

Rod-like bacteria.

Spirilla

Spiral-shaped bacteria.

Bacterial Growth Curve Steps of Growth Pattern:

Lag Phase: Bacteria adapt to their growth conditions.
Log (Exponential Phase): The number of cells and the rate of growth double.
Stationary Phase: Growth is limited by factors such as nutrient depletion, waste buildup, etc.
Death Phase: Bacteria die due to limited resources, unfavorable environmental conditions, etc.

Transformation

Bacteria take up genetic material from the environment.
Competent Bacteria are cells capable of undergoing transformation.

Transduction

Viruses transfer bacterial DNA.

Conjugation

Transfer of genetic material through a cytoplasmic bridge between bacteria.

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Description

Explore the fascinating world of microscopes in this quiz. Learn about different types of microscopes, including stereo, compound, and fluorescence microscopes. Understand key concepts such as fixation and staining that enhance specimen visibility.